Patent Publication Number: US-9428740-B2

Title: Lipase-containing polymeric coatings for the facilitated removal of fingerprints

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS 
     This patent application is a continuation of U.S. utility patent application Ser. No. 13/757,978 filed Feb. 4, 2013, which is a divisional of U.S. utility patent application Ser. No. 12/820,063, filed Jun. 21, 2010 (now U.S. Pat. No. 8,394,618), the entire contents of which are incorporated herein in their entirety. 
    
    
     FIELD OF THE INVENTION 
     The present invention relates generally to coating compositions including bioactive substances and methods of their use to facilitate removal of fingerprints. In specific embodiments, the invention relates to methods for fingerprint removal by incorporating lipase into or on polymer composite materials to degrade fingerprint components. 
     BACKGROUND OF THE INVENTION 
     Many consumer products such as cell phones, touch-screen displays, automobile door handles, etc., are subject to frequent contact with hands and fingers. As a result, the residue of fingerprints often leaves unpleasant marks on the surface deteriorating the cosmetic appearance of the products. 
     Prior art approaches aim to reduce the deposition of the fingerprint stains on a surface and facilitate its removal capitalize on the “lotus-effect” where hydrophobic, oleophobic and super-amphiphobic properties are conferred to the surface by polymeric coatings containing appropriate nanocomposites. An exemplary coating contains fluorine and silicon nanocomposites with good roll off properties and very high water and oil contact angles. When used on rough surfaces like sandblasted glass, nanocoatings may act as a filler to provide a fingerprint resistance. A drawback of these “passive” technologies is that they require water-rinsing to finally remove the fingerprints from the surface. In addition, these materials are not suitable for use in high gloss surfaces because the lotus-effect is based on surface roughness. 
     The photocatalyst Ti0 2  was used to promote active fingerprint decomposition of fingerprint stains in U.S. Pat. Appl. Publ. 2009/104086. A major drawback to this technology is its limitation to use on inorganic surfaces due to the oxidative impairment of the polymer coating by Ti0 2 . 
     Therefore, there is a need for new materials or coatings that can actively promote the removal of fingerprints on organic surfaces or in organic coatings and minimize the requirement for maintenance cleaning. 
     SUMMARY OF THE INVENTION 
     A composition and method for fingerprint removal from a substrate surface is provided. The method includes associating a lipase with a substrate or a coating such that the lipase is capable of enzymatically degrading a component of a fingerprint. 
     A method optionally includes heating the substrate or applying heat to the surface of the substrate. In some embodiments heating is at least 5 degrees Celsius above ambient temperature. The substrate or surface thereon is optionally heated to between 40 and 125 degrees Celsius. Heating is optionally continued for at least 30 minutes, illustratively for between 30 minutes to 6 hours. 
     The composition includes a substrate or coating containing a lipase. The composition optionally includes an organic crosslinkable or non-crosslinkable polymer resin. The resin optionally has a functional group of acetoacetate, acid, amine, carboxyl, epoxy, hydroxyl, isocyanate, silane, vinyl, or combinations thereof. Specific examples of resins include aminoplasts, melamine formaldehydes, carbamates, polyurethanes, polyacrylates, epoxies, polycarbonates, alkyds, vinyls, polyamides, polyolefins, phenolic resins, polyesters, polysiloxanes, or combinations thereof. In particular, a resin is optionally a hydroxyl-functionalized acrylate resin. 
     A substrate or coating has one or more associated lipase enzymes. A lipase is optionally lipoprotein lipase, acylglycerol lipase, hormone-sensitive lipase, phospholipase A1, phospholipase A2, phospholipase C, phospholipase D, phosphoinositide phospholipase C, a lysophospholipase, or a galactolipase. In particular embodiments, a lipase is a triacylglycerol lipase. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1  represents fingerprint removal on a substrate coated with a lipase containing coating; 
         FIG. 2  represents increased fingerprint removal rates on a plate coated with a lipase containing coating; 
         FIG. 3  represents similar removal rates of fingerprints from multiple sources. 
         FIG. 4  represents a schematic of an inventive method. 
     
    
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS 
     The following description of embodiment(s) of the invention is merely exemplary in nature and is in no way intended to limit the scope of the invention, its application, or uses, which may, of course, vary. The invention is described with relation to the non-limiting definitions and terminology included herein. These definitions and terminology are not designed to function as a limitation on the scope or practice of the invention but are presented for illustrative and descriptive purposes only. 
     The present invention is based on the catalytic activity of a lipase enzyme to selectively degrade and volatilize components of fingerprints, thus, promoting active fingerprint removal. Fingerprint stains typically include components of sweat gland secretion and sebum which includes lipids, wax, and cellular debris. Several of the substances of sebum are lipophilic and have low volatility such as squalene and wax esters. 
     The lipase immobilized in coatings or substrates catalyzes the hydrolysis, esterification, or transesterification of lipids including triacylglycerols, cholesterol esters, and other fingerprint components into smaller molecules. The smaller molecules may have higher volatility than their precursors and more easily vaporize at ambient or elevated temperatures thereby allowing for complete stain removal. Without being limited to one particular theory, it is believed that the resulting degradation products may have lower boiling points or reduced adhesion promoting increased vaporization either upon heating or incubation at ambient temperatures. Thus, the invention has utility as a composition and method for the active removal of fingerprints from surfaces. 
     The inventive methods and compositions are generally referred to herein as a lipase associated with a substrate for exemplary purposes only. One of ordinary skill in the art appreciates that the description is equally applicable to coatings either on a substrate or prior to application to a substrate. 
     An inventive method includes providing a substrate or a coating with a lipase or analogue thereof such that the lipase or analogue thereof is enzymatically active and capable of degrading one or more components of a fingerprint. In particular embodiments, a fingerprint is based on bioorganic matter such as that derived from the skin of a subject. 
     A fingerprint as defined herein is a bioorganic stain, mark, or residue left behind after an organism touches a substrate or coating. A fingerprint is not limited to marks or residue left behind after a substrate is touched by a finger. Other sources of bioorganic stains are illustratively, palms, toes, feet, face, any other skin surface area, hair, stains from fats used in cooking such as cis-fatty acids, or fatty acids from any other source. 
     A lipase is optionally a lipoprotein lipase, acylglycerol lipase such as triacylglycerol lipase, hormone-sensitive lipase, phospholipase A1, phospholipase A2, phospholipase C, phospholipase D, phosphoinositide phospholipase C, a lysophospholipase, a galactolipase, or combinations or analogues thereof. An analogue of a lipase is optionally a fragment of a lipase. An analogue of a lipase is a polypeptide that has some level of activity toward a natural or synthetic substrate of a lipase. An analogue optionally has between 0.1% and 200% the activity of a wild-type lipase. 
     Specific examples of lipase include Lipase AP4, Lipase AP6, Lipase AP12, Lipase M-AP5, Lipase M-AP10 and Lipase M-AP20 (manufactured by Amano Pharmaceutical Co., Ltd.), Lipase Saiken (manufactured by Osaka Saikin Kenkyusho), Lipase MY (manufactured by Meito Sangyo), or Lipase B ( Candida antarctica  or  Candida rugosa ). It is also possible to use multiple enzyme preparations having lipase activity. For example, mixed digestive enzyme preparations are operable such as Biodiastase, Biodiastase 500, Biodiastase 700, Biodiastase 1000, Biodiastase 2000, Pancreatin, Pancreatic Digestive Enzyme TA and Pancreatic Digestive Enzyme 8AP (manufactured by Amano Pharmaceutical Co., Ltd.), Biotamylase, Biotamyolase S, Biotalase A-1000, Biotalase P-1000 and Denapsin 10 (manufactured by Nagase Seikagaku Kogyo), Cellulosin AP and Prolicin (manufactured by Ueda Kagaku), Takadiastase (manufactured by Sankyo), Sumizyme (manufactured by Shin Nippon Kagaku Kogyo) and Biotamylase (manufactured by Nagase Sangyo). 
     A lipase is optionally derived from  Acinetobacter, Aedes aegypti, Anguilla japonica, Antrodia cinnamomea, Arabidopsis rosette, Arabidopsis thaliana, Arxula adeninivorans, Aspergillus niger, Aspergillus oryzae, Aspergillus tamarii, Aureobasidium pullulans, Avena sativa, Bacillus licheniformis, Bacillus sphaericus, Bacillus stearothermophilus, Bacillus subtilis, Bacillus thermocatenulatus, Bacillus thermoleovorans, Bombyx mandarina, Bombyx mori, Bos Taurus, Brassica napus, Brassica rapa, Burkholderia cepacia, Caenorhabditis elegans, Candida albicans, Candida antarctica, Candida deformans, Candida parapsilosis, Candida rugosa, Candida thermophile, Canis domesticus, Chenopodium rubrum, Clostridium beijerinckii, Clostridium botulinum, Clostridium novyi, Danio rerio, Galactomyces geotrichum, Gallus gallus, Geobacillus, Gibberella zeae, Gossypium hirsutum, Homo sapiens, Kurtzmanomyces  sp.,  Leishmania infantum, Lycopersicon esculentum  L,  Malassezia furfur, Methanosarcina acetivorans, Mus musculus, Mus spretus, Mycobacterium tuberculosis, Mycoplasma hyopneumoniae, Myxococcus xanthus, Neosartorya fischeri, Oryctolagus cuniculus, Oryza sativa, Penicillium cye/opium, Phlebotomus papatasi, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas fragi, Pseudomonas  sp,  Rattus norvegicus, Rhizomucor miehei, Rhizopus oryzae, Rhizopus stolonifer, Ricinus communis, Samia Cynthia ricini, Schizosaccharomyces pombe, Serratia marcescens, Spermophilus tridecemlineatus, Staphylococcus simulans, Staphylococcus xylosus, Sulfolobus solfataricus, Sus scrofa, Thermomyces lanuginosus, Trichomonas vaginalis, Vibrio harveyi, Xenopus laevis, Yarrowia lipolytica , a combination thereof, or a derivative thereof. It is appreciated that lipases derived from other organisms are similarly operable and are within the scope of the present invention. 
     A lipase is a “peptide,” “polypeptide,” and “protein” that are used herein synonymously and are intended to mean a natural or synthetic compound containing two or more amino acids having some level of activity toward a natural or synthetic substrate of a wild-type lipase. A wild-type lipase is a lipase that has an amino acid sequence identical to that found in an organism in nature. An illustrative example of a wild-type lipase is that found at GenBank Accession No. ACL68189 and SEQ ID NO: 1. An exemplary nucleotide sequence encoding a wild-type lipase is found at Accession No. FJ536288 and SEQ ID NO: 2. One of skill in the art recognizes how to modify a nucleotide sequence to alter or create a protein sequence. 
     
       
         
           
               
               
            
               
                   
                 (SEQ ID NO: 1) 
               
               
                   
                 MFLRREFGAV AALSVLAHAA PAPAPMQRRD ISSTVLDNID 
               
               
                   
                   
               
               
                   
                 LFAQYSAAAY CSSNIESTGT TLTCDVGNCP LVEAAGATTI 
               
               
                   
                   
               
               
                   
                 DEFDDSSSYG DPTGFIAVDP TNELIVLSFR GSSDLSNWIA 
               
               
                   
                   
               
               
                   
                 DLDFGLTSVS SICDGCEMHK GFYEAWEVIA DTITSKVEAA 
               
               
                   
                   
               
               
                   
                 VSSYPDYTLV FTGHSYGAAL AAVAATVLRN AGYTLDLYNF 
               
               
                   
                   
               
               
                   
                 GQPRIGNLAL ADYITDQNMG SNYRVTHTDD IVPKLPPELL 
               
               
                   
                   
               
               
                   
                 GYHHFSPEYW ITSGNDVTVT TSDVTEVVGV DSTDGNDGTL 
               
               
                   
                   
               
               
                   
                 LDSTTAHRWY TIYISECS 
               
               
                   
                   
               
               
                   
                 (SEQ ID NO: 2) 
               
               
                   
                 atgtttctcc gcagggaatt tggggctgtt gcagccctat 
               
               
                   
                   
               
               
                   
                 ctgtgctggc ccatgctgct cccgcacctg ctccgatgca 
               
               
                   
                   
               
               
                   
                 gcgtagagac atctcctcta ccgtcttgga caatatcgac 
               
               
                   
                   
               
               
                   
                 ctcttcgccc aatacagtgc agcagcttac tgctcctcca 
               
               
                   
                   
               
               
                   
                 acatcgagtc caccggcacg actctgacct gcgacgtagg 
               
               
                   
                   
               
               
                   
                 caattgccct ctcgtcgagg cagccggtgc cacgaccatc 
               
               
                   
                   
               
               
                   
                 gatgagtttg acgacagcag cagctacggc gacccgacgg 
               
               
                   
                   
               
               
                   
                 ggttcatcgc cgttgacccg acgaacgagt tgatcgttct 
               
               
                   
                   
               
               
                   
                 gtctttccgg ggtagttccg acctctcgaa ctggattgcc 
               
               
                   
                   
               
               
                   
                 gacctagact tcggcctcac ctccgtaagc agcatctgtg 
               
               
                   
                   
               
               
                   
                 atggctgtga gatgcacaag ggcttctatg aggcctggga 
               
               
                   
                   
               
               
                   
                 agtcattgcc gacaccatca catccaaggt ggaggccgct 
               
               
                   
                   
               
               
                   
                 gtctccagct atccggacta caccctcgtg ttcaccggac 
               
               
                   
                   
               
               
                   
                 acagctacgg cgctgcattg gcggctgtcg cggccaccgt 
               
               
                   
                   
               
               
                   
                 actccgcaac gccggataca ctcttgacct gtacaacttc 
               
               
                   
                   
               
               
                   
                 ggccagcccc gtatcggcaa ccttgcttta gctgactaca 
               
               
                   
                   
               
               
                   
                 tcaccgacca aaacatgggc agcaactacc gcgtcacgca 
               
               
                   
                   
               
               
                   
                 caccgacgac atcgtgccta agctgcctcc ggagctgctg 
               
               
                   
                   
               
               
                   
                 ggctaccacc acttcagtcc ggagtactgg atcaccagcg 
               
               
                   
                   
               
               
                   
                 gtaatgatgt gacggtgact acgtcggacg tgaccgaggt 
               
               
                   
                   
               
               
                   
                 tgtgggggtg gattcgacgg atgggaatga cggcacgctg 
               
               
                   
                   
               
               
                   
                 cttgacagta cgactgccca tcggtggtac acgatctaca 
               
               
                   
                   
               
               
                   
                 ttagtgaatg ctcgtag 
               
            
           
         
       
     
     Lipase activity is illustratively defined in units/gram. 1 unit illustratively corresponds to the amount of enzyme that hydrolyzes 1 μmol acetic acid per minute at pH 7.4 and 40° C. using the substrate triacetin (Sigma-Aldrich, St. Louis, Mo., Product No. 90240). The lipase of SEQ ID NO: 1 has an activity of approximately 200 units/gram. 
     Methods of screening for lipase activity are known and standard in the art. Illustratively, screening for lipase activity in a lipase protein or analogue thereof illustratively includes contacting a lipase or analogue thereof with a natural or synthetic substrate of a lipase and measuring the enzymatic cleavage of the substrate. Illustrative substrates for this purpose include tributyrin and triacetin both of which are cleaved by a triacylglycerol lipase to liberate butyric acid or acetic acid respectively that is readily measured by techniques known in the art. 
     Amino acids present in a lipase or analogue thereof illustratively include the common amino acids alanine, cysteine, aspartic acid, glutamic acid, phenylalanine, glycine, histidine, isoleucine, lysine, leucine, methionine, asparagine, proline, glutamine, arginine, serine, threonine, valine, tryptophan, and tyrosine; as well as less common naturally occurring amino acids, modified amino acids or synthetic compounds, such as alpha-asparagine, 2-aminobutanoic acid or 2-aminobutyric acid, 4-aminobutyric acid, 2-aminocapric acid (2-aminodecanoic acid), 6-aminocaproic acid, alpha-glutamine, 2-aminoheptanoic acid, 6-aminohexanoic acid, alpha-aminoisobutryic acid (2-aminoalanine), 3-aminoisobutyric acid, beta-alanine, allo-hydroxylysine, allo-isoleucine, 4-amino-7-methylheptanoic acid, 4-amino-5-phenylpentanoic acid, 2-aminopimelic acid, gamma-amino-beta-hydroxybenzenepentanoic acid, 2-aminosuberic acid, 2-carboxyazetidine, beta-alanine, beta-aspartic acid, biphenylalanine, 3,6-diaminohexanoic acid, butanoic acid, cyclobutyl alanine, cyclohexylalanine, cyclohexylglycine, N5-aminocarbonylornithine, cyclopentyl alanine, cyclopropyl alanine, 3-sulfoalanine, 2,4-diaminobutanoic acid, diaminopropionic acid, 2,4-diaminobutyric acid, diphenyl alanine, N,N-dimethylglycine, diaminopimelic acid, 2,3-diaminopropanoic acid, S-ethylthiocysteine, N-ethylasparagine, N-ethylglycine, 4-aza-phenylalanine, 4-fluoro-phenylalanine, gamma-glutamic acid, gamma-carboxyglutamic acid, hydroxyacetic acid, pyroglutamic acid, homoarginine, homocysteic acid, homocysteine, homohistidine, 2-hydroxyisovaleric acid, homophenylalanine, homoleucine, homoproline, homoserine, homoserine, 2-hydroxypentanoic acid, 5-hydroxylysine, 4-hydroxyproline, 2-carboxyoctahydroindole, 3-carboxyisoquinoline, isovaline, 2-hydroxypropanoic acid (lactic acid), mercaptoacetic acid, mercaptobutanoic acid, sarcosine, 4-methyl-3-hydroxyproline, mercaptopropanoic acid, norleucine, nipecotic acid, nortyrosine, norvaline, omega-amino acid, ornithine, penicillamine (3-mercaptovaline), 2-phenylglycine, 2-carboxypiperidine, sarcosine (N-methylglycine), 2-amino-3-(4-sulfophenyl)propionic acid, 1-amino-1-carboxycyclopentane, 3-thienylalanine, epsilon-N-trimethyllysine, 3-thiazolylalanine, thiazolidine 4-carboxylic acid, alpha-amino-2,4-dioxopyrimidinepropanoic acid, and 2-naphthylalanine. A lipase includes peptides having between 2 and about 1000 amino acids or having a molecular weight in the range of about 150-350,000 Daltons. 
     A lipase is obtained by any of various methods known in the art illustratively including isolation from a cell or organism, chemical synthesis, expression of a nucleic acid sequence, and partial hydrolysis of proteins. Chemical methods of peptide synthesis are known in the art and include solid phase peptide synthesis and solution phase peptide synthesis or by the method of Hackeng, T M, et al.,  Proc Natl Acad Sci USA,  1997; 94(15):7845-50, the contents of which are incorporated herein by reference. A lipase included in an inventive composition may be a naturally occurring or non-naturally occurring protein. The term “naturally occurring” refers to a protein endogenous to a cell, tissue or organism and includes allelic variations. A non-naturally occurring peptide is synthetic or produced apart from its naturally associated organism or is modified and is not found in an unmodified cell, tissue, or organism. 
     Modifications and changes can be made in the structure of a lipase and still obtain a molecule having similar characteristics as lipase (e.g., a conservative amino acid substitution). For example, certain amino acids can be substituted for other amino acids in a sequence without appreciable loss of activity or optionally to reduce or increase the activity of an unmodified lipase. Because it is the interactive capacity and nature of a polypeptide that defines that polypeptide&#39;s biological functional activity, certain amino acid sequence substitutions can be made in a polypeptide sequence and nevertheless obtain a polypeptide with like or other desired properties. 
     In making such changes, the hydropathic index of amino acids can be considered. The importance of the hydropathic amino acid index in conferring interactive biologic function on a polypeptide is generally understood in the art. It is known that certain amino acids can be substituted for other amino acids having a similar hydropathic index or score and still result in a polypeptide with similar biological activity. Each amino acid has been assigned a hydropathic index on the basis of its hydrophobicity and charge characteristics. Those indices are: isoleucine (+4.5); valine (+4.2); leucine (+3.8); phenylalanine (+2.8); cysteine/cysteine (+2.5); methionine (+1.9); alanine (+1.8); glycine (−0.4); threonine (−0.7); serine (−0.8); tryptophan (−0.9); tyrosine (−1.3); proline (−1.6); histidine (−3.2); glutamate (−3.5); glutamine (−3.5); aspartate (−3.5); asparagine (−3.5); lysine (−3.9); and arginine (−4.5). 
     It is believed that the relative hydropathic character of the amino acid determines the secondary structure of the resultant polypeptide, which in turn defines the interaction of the polypeptide with other molecules, such as enzymes, substrates, receptors, antibodies, antigens, and the like. It is known in the art that an amino acid can be substituted by another amino acid having a similar hydropathic index and still obtain a functionally equivalent polypeptide. In such changes, the substitution using amino acids whose hydropathic indices are within ±2, those within ±1, and those within ±0.5 are optionally used. 
     Substitution of like amino acids can also be made on the basis of hydrophilicity. The following hydrophilicity values have been assigned to amino acid residues: arginine (+3.0); lysine (+3.0); aspartate (+3.0±1); glutamate (+3.0±1); serine (+0.3); asparagine (+0.2); glutamine (+0.2); glycine (0); proline (−0.5±1); threonine (−0.4); alanine (−0.5); histidine (−0.5); cysteine (−1.0); methionine (−1.3); valine (−1.5); leucine (−1.8); isoleucine (−1.8); tyrosine (−2.3); phenylalanine (−2.5); tryptophan (−3.4). It is understood that an amino acid can be substituted for another having a similar hydrophilicity value and still obtain an enzymatically equivalent polypeptide. In such changes, the substitution of amino acids whose hydrophilicity values are within ±2, those within ±1, and those within ±0.5 are optionally used. 
     Amino acid substitutions are optionally based on the relative similarity of the amino acid side-chain substituents, for example, their hydrophobicity, hydrophilicity, charge, size, and the like. Exemplary substitutions that take various of the foregoing characteristics into consideration are well known to those of skill in the art and include (original residue: exemplary substitution): (Ala: Gly, Ser), (Arg: Lys), (Asn: Gln, His), (Asp: Glu, Cys, Ser), (Gln: Asn), (Glu: Asp), (Gly: Ala), (His: Asn, Gln), (Ile: Leu, Val), (Leu: Ile, Val), (Lys: Arg), (Met: Leu, Tyr), (Ser: Thr), (Thr: Ser), (Tip: Tyr), (Tyr: Trp, Phe), and (Val: Ile, Leu). Embodiments of this disclosure, thus, contemplate functional or biological equivalents of a polypeptide as set forth above. In particular, embodiments of the polypeptides can include analogues having about 50%, 60%, 70%, 80%, 90%, 95%, or 99% sequence identity to a wild-type lipase. 
     It is further appreciated that the above characteristics are optionally taken into account when producing a lipase with reduced or increased enzymatic activity. Illustratively, substitutions in a substrate binding site, exosite, cofactor binding site, catalytic site, or other site in a lipase protein may alter the activity of the enzyme toward a substrate. In considering such substitutions the sequences of other known naturally occurring or non-naturally occurring lipases may be taken into account. Illustratively, the N291S substitution in lipoprotein lipase reduces its enzymatic activity by 30-50 percent. Busca, R. et al,  FEBS Lett.,  1995; 367:257-262, the contents of which are incorporated herein by reference. Illustratively substitution at amino acid 95 in  Rhizomucor miehei  lipase can produce a 2-3 fold increase in activity toward particular substrates. Similarly substitution at amino acid 94 can produce as much as 6 times the activity of wild-type. Oh, S, et al, Biotech. Lett., 2001; 23: 563-568, the contents of which are incorporated herein by reference. Other substitutions at this or other sites may similarly affect enzymatic activity. 
     A lipase protein is illustratively recombinant. Methods of cloning, synthesizing or otherwise obtaining nucleic acid sequences encoding a lipase are known and standard in the art that are equally applicable to lipase. Similarly, methods of cell transfection and protein expression are similarly known in the art and are applicable herein. Such methods are illustratively disclosed in Molecular Cloning: A Laboratory Manual, 3rd ed., vol. 1-3, ed. Sambrook et al., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 2001; Current Protocols in Molecular Biology, ed. Ausubel et al., Greene Publishing and Wiley-Interscience, New York, 1992 (with periodic updates); and Short Protocols in Molecular Biology, ed. Ausubel et al., 52 ed., Wiley-Interscience, New York, 2002, the contents of each of which are incorporated herein by reference. 
     A lipase may be coexpressed with associated tags, modifications, other proteins such as a fusion peptide, or other modifications or combinations recognized in the art. Illustrative tags include 6× His, FLAG, biotin, ubiquitin, SUMO, or other tag known in the art. A tag is illustratively cleavable such as by linking to lipase or an associated protein via an enzyme cleavage sequence that is cleavable by an enzyme known in the art illustratively including Factor Xa, thrombin, SUMOstar protein as obtainable from Lifesensors, Inc., Malvern, Pa., or trypsin. It is further appreciated that chemical cleavage is similarly operable with an appropriate cleavable linker. 
     Protein expression is illustratively accomplished from transcription of a lipase nucleic acid sequence, translation of RNA transcribed from the lipase nucleic acid sequence or analogues thereof. Protein expression is optionally performed in a cell based system such as in  E. coli , Hela cells, or Chinese hamster ovary cells. It is appreciated that cell-free expression systems are similarly operable. 
     It is recognized that numerous analogues of lipase are operable and within the scope of the present invention including amino acid substitutions, alterations, modifications, or other amino acid changes that increase, decrease, or do not alter the function of the lipase protein sequence. Several post-translational modifications are similarly envisioned as within the scope of the present invention illustratively including incorporation of a non-naturally occurring amino acid, phosphorylation, glycosylation, and addition of pendent groups such as biotinylation, fluorophores, lumiphores, radioactive groups, antigens, or other molecules. 
     An inventive method uses an inventive composition that is one or more lipases incorporated into a substrate itself or into a coating on the substrate. The lipase enzyme is optionally non-covalently associated and/or covalently attached to the substrate or coating material or is otherwise associated therewith such as by bonding to the surface or by intermixing with the substrate/coating material during manufacture such as to produce entrapped lipase. In some embodiments the lipase is covalently attached to the substrate or coating material either by direct covalent interaction between the lipase and one or more components of the substrate or coating material or by association via a link moiety such as that described in U.S. Pat. App. Publ. No. 2008/0119381, the contents of which are incorporated herein by reference. 
     There are several ways to associate lipase with a substrate or coating. One of which involves the application of covalent bonds. Specifically, free amine groups of the lipase may be covalently bound to an active group of the substrate. Such active groups include alcohol, thiol, aldehyde, carboxylic acid, anhydride, epoxy, ester, or any combination thereof. This method of incorporating lipase delivers unique advantages. First, the covalent bonds tether the lipases permanently to the substrate and thus places them as an integral part of the final composition with much less, if any at all, leakage of the lipase. Second, the covalent bonds provide extended enzyme lifetime. Over time proteins typically lose activity because of the unfolding of their polypeptide chains. Chemical binding such as covalent bonding effectively restricts such unfolding, and thus improves the protein life. The life of a protein is typically determined by comparing the amount of activity reduction of a protein that is free or being physically adsorbed with that of a protein covalently-immobilized over a period of time. 
     Lipases are optionally uniformly dispersed throughout the substrate network to create a substantially homogenous protein platform. In so doing, lipases may be first modified with polymerizable groups. The modified lipases may be solubilized into organic solvents in the presence of surfactant, and thus engage the subsequent polymerization with monomers such as methyl methacrylate (MMA) or styrene in the organic solution. The resultant composition includes lipase molecules homogeneously dispersed throughout the network. 
     Lipases are optionally attached to surfaces of a substrate. An attachment of lipases corresponding to approximately 100% surface coverage was achieved with polystyrene particles with diameters range from 100 to 1000 nm. 
     Chemical methods of lipase attachment to materials will naturally vary depending on the functional groups present in the lipase and in the material components. Many such methods exist. For example, methods of attaching proteins (such as enzymes) to other substances are described in O&#39;Sullivan et al,  Methods in Enzymology,  1981; 73:147-166 and Erlanger,  Methods in Enzymology,  1980; 70:85-104, each of which are herein incorporated herein by reference. 
     Lipases are optionally present in a coating that is layered upon a substrate wherein the lipase is optionally entrapped in the coating material, admixed therewith, modified and integrated into the coating material or layered upon a coating similar to the mechanisms described for interactions between a lipase and substrate material. 
     Materials operable for interactions with a lipase to form an active substrate or coating illustratively include organic polymeric materials. The combination of these materials and a lipase form a protein-polymer composite material that is used as a substrate material or a coating. 
     Methods of preparing protein-polymer composite materials illustratively include use of aqueous solutions of lipase and non-aqueous organic solvent-borne polymers to produce bioactive organic solvent-borne protein-polymer composite materials. 
     Methods of preparing protein-polymer composite materials are illustratively characterized by dispersion of lipase in solvent-borne resin prior to curing and in the composite materials. Lipases are optionally dispersed in the protein-polymer composite material such that the lipases are unassociated with other bioactive proteins and/or form relatively small particles of associated proteins. Illustratively, the average particle size of lipase particles in the protein-polymer composite material is less than 10 μm (average diameter) such as in the range of 1 nm to 10 μm, inclusive. 
     Curable protein-polymer compositions are optionally two-component solvent-borne (2K SB) compositions. Optionally, one component systems (1K) are similarly operable. Illustratively, a lipase is entrapped in a coating material such as a latex or enamel paint, varnish, polyurethane gels, or other coating materials. Illustrative examples of incorporating enzymes into paints are presented in U.S. Pat. No. 5,998,200, the contents of which are incorporated herein by reference. 
     In two-component (2K) systems the two components are optionally mixed shortly before use, for instance, application of the curable protein-polymer composition to a substrate to form a lipase containing coating such as a bioactive clear coat. Generally described, the first component contains a crosslinkable polymer resin and the second component contains a crosslinker. Thus, the emulsion is a first component containing a crosslinkable resin and the crosslinker is a second component, mixed together to produce the curable protein-polymer composition. 
     A polymer resin included in methods and compositions of the present invention can be any film-forming polymer useful in coating or substrate compositions, illustratively clear coat compositions. Such polymers illustratively include, aminoplasts, melamine formaldehydes, carbamates, polyurethanes, polyacrylates, epoxies, polycarbonates, alkyds, vinyls, polyamides, polyolefins, phenolic resins, polyesters, polysiloxanes; and combinations of any of these or other polymers. 
     In particular embodiments, a polymer resin is crosslinkable. Illustratively, a crosslinkable polymer has a functional group characteristic of a crosslinkable polymer. Examples of such functional groups illustratively include acetoacetate, acid, amine, carboxyl, epoxy, hydroxyl, isocyanate, silane, vinyl, other operable functional groups, and combinations thereof. 
     Examples of organic crosslinkable polymer resins include aminoplasts, melamine formaldehydes, carbamates, polyurethanes, polyacrylates, epoxies, polycarbonates, alkyds, vinyls, polyamides, polyolefins, phenolic resins, polyesters, polysiloxanes, or combinations thereof. 
     A crosslinking agent is optionally included in the composition. The particular crosslinker selected depends on the particular polymer resin used. Non-limiting examples of crosslinkers include compounds having functional groups such as isocyanate functional groups, epoxy functional groups, aldehyde functional groups, and acid functionality. 
     In particular embodiments of protein-polyurethane composite materials, a polymer resin is a hydroxyl-functional acrylic polymer and the crosslinker is a polyisocyanate. 
     A polyisocyanate, optionally a diisocyanate, is a crosslinker reacted with the hydroxyl-functional acrylic polymer according to embodiments of the present invention. Aliphatic polyisocyanates are preferred polyisocyanates used in processes for making protein-polymer composite materials for clearcoat applications such as in automotive clearcoat applications. Non-limiting examples of aliphatic polyisocyanates include 1,4-butylene diisocyanate, 1,4-cyclohexane diisocyanate, 1,2-diisocyanatopropane, 1,3-diisocyanatopropane, ethylene diisocyanate, lysine diisocyanate, 1,4-methylene bis (cyclohexyl isocyanate), diphenylmethane 4,4′-diisocyanate, an isocyanurate of diphenylmethane 4,4′-diisocyanate, methylenebis-4,4′-isocyanatocyclohexane, 1,6-hexamethylene diisocyanate, an isocyanurate of 1,6-hexamethylene diisocyanate, isophorone diisocyanate, an isocyanurate of isophorone diisocyanate, p-phenylene diisocyanate, toluene diisocyanate, an isocyanurate of toluene diisocyanate, triphenylmethane 4,4′,4″-triisocyanate, tetramethyl xylene diisocyanate, and meta-xylene diisocyanate. 
     Curing modalities are those typically used for conventional curable polymer compositions. 
     Lipase-polymer composite materials used in embodiments of the present invention are optionally thermoset protein-polymer composite materials. For example, a substrate or coating material is optionally cured by thermal curing. A thermal polymerization initiator is optionally included in a curable composition. Thermal polymerization initiators illustratively include free radical initiators such as organic peroxides and azo compounds. Examples of organic peroxide thermal initiators illustratively include benzoyl peroxide, dicumylperoxide, and lauryl peroxide. An exemplary azo compound thermal initiator is 2,2′-azobisisobutyronitrile. 
     Conventional curing temperatures and curing times can be used in processes according to embodiments of the present invention. For example, the curing time at specific temperatures, or under particular curing conditions, is determined by the criteria that the cross-linker functional groups are reduced to less than 5% of the total present before curing. Cross-linker functional groups can be quantitatively characterized by FT-IR or other suitable method. For example, the curing time at specific temperatures, or under particular curing conditions, for a polyurethane protein-polymer composite of the present invention can be determined by the criteria that the cross-linker functional group NCO is reduced to less than 5% of the total present before curing. The NCO group can be quantitatively characterized by FT-IR. Additional methods for assessing the extent of curing for particular resins are well-known in the art. Illustratively, curing may include evaporation of a solvent or by exposure to actinic radiation, such as ultraviolet, electron beam, microwave, visible, infrared, or gamma radiation. 
     One or more additives are optionally included for modifying the properties of the lipase-polymer composite material and/or the admixture of organic solvent and polymer resin, the aqueous lipase solution, the emulsion, and/or the curable composition. Illustrative examples of such additives include a UV absorbing agent, a plasticizer, a wetting agent, a preservative, a surfactant, a lubricant, a pigment, a filler, and an additive such as an additive to increase sag resistance. 
     A substrate or coating including a lipase is illustratively an admixture of a polymer resin, a surfactant and a non-aqueous organic solvent, mixed to produce an emulsion. The term “surfactant” refers to a surface active agent that reduces the surface tension of a liquid in which it is dissolved, or that reduces interfacial tension between two liquids or between a liquid and a solid. 
     Surfactants used can be of any variety including amphoteric, silicone-based, fluorosurfactants, anionic, cationic and nonionic such as described in K. R. Lange, Surfactants: A Practical Handbook, Hanser Gardner Publications, 1999; and R. M. Hill, Silicone Surfactants, CRC Press, 1999. Examples of anionic surfactants include alkyl sulfonates, alkylaryl sulfonates, alkyl sulfates, alkyl and alkylaryl disulfonates, sulfonated fatty acids, sulfates of hydroxyalkanols, sulfosuccinic acid esters, sulfates and sulfonates of polyethoxylated alkanols and alkylphenols. Examples of cationic surfactants include quaternary surfactants and amineoxides. Examples of nonionic surfactants include alkoxylates, alkanolamides, fatty acid esters of sorbitol or manitol, and alkyl glucamides. Examples of silicone-based surfactants include siloxane polyoxyalkylene copolymers. 
     When a surface which is optionally a substrate or a coated substrate, is contacted with a fingerprint, the lipase enzyme or combinations of enzymes contact the fingerprint, or components thereof. The contacting allows the enzymatic activity of the substrate or coating to interact with and enzymatically alter the components of the fingerprint improving their removal from the substrate or coating. 
     It is appreciated that the inventive methods of facilitating fingerprint removal will function at any temperature whereby the lipase is active. Optionally, the inventive method is performed at 4° C. Optionally, an inventive method is performed at 25° C. Optionally, an inventive process is performed at ambient temperature. Some embodiments are performed between 40° C. and 120° C. 
     An inventive method optionally includes heating the substrate itself or applying heat to the surface of the substrate. Heating is defined as increasing the surface temperature of a substrate/coating to a level higher than the ambient temperature. In some embodiments heating is raising the surface temperature by at least 5° C. Heating is optionally raised by exposure to sunlight or other heat source. The surface temperature is optionally raised to such a level that the breakdown products volatilize to the point of no visual material remaining on the substrate within 24 hours. Optionally, the temperature is raised to such a level that the breakdown products are removed to the point of no visual material remaining on the substrate within 0.5 to 3 hours, inclusive. Optionally, the substrate surface temperature is increased to between 40° C. and 125° C., inclusive. 
     Heat is optionally applied continuously or intermittently. Heat is optionally applied until the breakdown products volatilize to the point of no visual material remaining on the substrate. Optionally, heat is applied for at least 30 minutes. In some embodiments, heat is applied for between 30 minutes to 6 hours, inclusive. 
     The presence of lipase combined with the material of a substrate or a coating on a substrate, optionally, with applied heat, breaks down fingerprint stains for facilitated fingerprint removal. 
     Various aspects of the present invention are illustrated by the following non-limiting examples. The examples are for illustrative purposes and are not a limitation on any practice of the present invention. It will be understood that variations and modifications can be made without departing from the spirit and scope of the invention. 
     Example 1 
     Production of lipase containing material operable for coating a substrate. 
     Materials: Polyacrylate resin Desmophen A870 BA, and the hexamethylene diisocyanate (HDI) based polyfunctional aliphatic polyisocyanate resin Desmodur N 3600, the polyester resin Bayhydrol XP 7093, and Bayhydur 302 are obtained from Bayer Corp. (Pittsburgh, Pa.). The surfactant BYK-333 is obtained from BYK-Chemie (Wallingford, Conn.). α-Amylase KLEISTASE SD80 from  Bacillus subtilis  (EC 3.2.1.1), lipase PS from  Burkholderia cepacia , and Lipase AP12 from  Aspergillus niger  are obtained from Amano Enzyme Inc. (Nagoya, Japan). Butyl acetate, Bradford reagent, bovine serum albumin (BSA) from bovine serum, starch from potatoes, starch from wheat, maltose, sodium potassium tartrate, 3,5-dinitrosalicylic acid, Na 2 (PO 4 ), NaCl, K 2 (PO 4 ), casein, trichloroacetic acid, Folin &amp; Ciocalteu&#39;s phenol reagent, Na 2 (CO 3 ), sodium acetate, calcium acetate, tyrosine, p-nitrophenyl palmitate, ethanol, iodine, glucose, maltose, maltotriose, maltohexose, dextrin with different molecular weight (10 kDa, 40 kDa) were obtained from Sigma Chemical Co., St. Louis, Mo., U.S.A. Aluminum panels and 8-path wet film applicators are obtained from Paul N. Gardner Company, Inc. (Pompano Beach, Fla.). 
     Enzyme preparation: Lipase AP12 is subjected to purification from a 150 mL solution containing 7.5 g crude lipase powder in DI water at a protein concentration of about 5 mg/mL (determined by the Bradford method). All solutions are kept on ice. Ultrafiltration is performed using a 150 mL Amicon cell (Millipore, Billerica, Mass.) at a pressure of 55 psi using a 30 kDa cut-off membrane. Ultrafiltration is repeated  3  times by adding DI water to the remaining concentrated solution to a volume of 150 mL after each run. The final remaining purified protein solution is used to prepare coatings typically at a lipase concentration of 200 mg/mL. 
     Incorporation of lipase in to form protein-polymer coating: Solvent-borne two-component polyurethane (PU) is formed by the polymerization of Desmophen A 870 BA and Desmodur N 3600 at a weight ratio of 2.6:1 of Desmophen A 870 BA to Desmodur N 3600. Typically, 2.1 g of Desmophen A 870 BA (solid content of 70 wt %), 0.0167 g of BYK-333 dissolved in 100 μl of n-butanol, and 0.5 mL of butyl acetate are mixed in a 20 mL vial. 600 μl of 200 mg/mL purified lipase enzyme solution is added and mixed for 1 minute to form a white emulsion. Subsequently, 0.8 g of Desmodur N 3600 is added and mixed manually for 1 minute. The resulting curable protein-polymer solution is then applied to aluminum testing panels via drawdown using an 8-path applicator. The resulting coating is cured at 80° C. for 24 hours. 
     Example 2 
     Fingerprint removal: The lipase containing coated panels of Example 1 are loaded with human fingerprints after touching the skin of the face or forearms. Fingerprinted panels are incubated at room temperature for at least 24 hours. A control panel is coated with the coating of Example 1 that is free of enzyme. After this first incubation period, the coated substrate is incubated in an oven at a temperature of 65° C. or higher for 1 to 6 hours. 
       FIG. 1  demonstrates that incubation of the enzyme coated panels at 65° C. for two hours facilitates complete removal of fingerprints. (B: control; L: lipase; LA: combined lipase and amylase in coating.) 
     Example 3 
     The rate of fingerprint removal is faster in an enzyme containing substrate than control.  FIG. 2  demonstrates aluminum plates coated with control or enzyme containing protein-polymer coatings as described in Example 1 where a fingerprint is loaded at the interface between the two coatings as described in Example 2.  FIG. 2A  demonstrates the remaining fingerprint after incubation at ambient temperature for 3 days. The plates are subsequently incubated for 2.5 hours at 65° C. and photographs are taken at various intervals as illustrated in  FIG. 2B .  FIG. 3  demonstrates that the increased rate of fingerprint removal is independent of the source of the fingerprints. 
     Various modifications of the present invention, in addition to those shown and described herein, will be apparent to those skilled in the art of the above description. Such modifications are also intended to fall within the scope of the appended claims. 
     It is appreciated that all reagents are obtainable by sources known in the art unless otherwise specified. Methods of nucleotide amplification, cell transfection, and protein expression and purification are similarly within the level of skill in the art. 
     Patents and publications mentioned in the specification are indicative of the levels of those skilled in the art to which the invention pertains. These patents and publications are incorporated herein by reference to the same extent as if each individual application or publication was specifically and individually incorporated herein by reference. 
     The foregoing description is illustrative of particular embodiments of the invention, but is not meant to be a limitation upon the practice thereof. The following claims, including all equivalents thereof, are intended to define the scope of the invention.