gebhart/patrepeval huggingface dataset:
Survey of Protein Engineering Strategies (2011)

Protein engineering is altering the structure of a protein to improve or change its properties. This unit summarizes concepts for protein engineering using rational design, directed evolution, and combinations of them. Different strategies are presented for identifying the best mutagenesis method, how to identify desired variants by screening or selection, and examples for successful applications are given. This should enable researchers to choose the most promising tools to solve their protein engineering challenges.
The viruses in all of us: characteristics and biological significance of human endogenous retrovirus sequences. (1996)

Human endogenous retroviruses (HERVs) are very likely footprints of ancient germ-cell infections. HERV sequences encompass about 1% of the human genome. HERVs have retained the potential of other retroelements to retrotranspose and thus to change genomic structure and function. The genomes of almost all HERV families are highly defective. Recent progress has allowed the identification of the biologically most active family, HTDV/HERV-K, which codes for viral proteins and particles and is highly expressed in germ-cell tumors. The demonstrable and potential roles of HTDV/HERV-K as well as of other human elements in disease and in maintaining genome plasticity are illustrated.
Site Specific Mutation of the Zic2 Locus by Microinjection of TALEN mRNA in Mouse CD1, C3H and C57BL/6J Oocytes (2013)

Transcription Activator-Like Effector Nucleases (TALENs) consist of a nuclease domain fused to a DNA binding domain which is engineered to bind to any genomic sequence. These chimeric enzymes can be used to introduce a double strand break at a specific genomic site which then can become the substrate for error-prone non-homologous end joining (NHEJ), generating mutations at the site of cleavage. In this report we investigate the feasibility of achieving targeted mutagenesis by microinjection of TALEN mRNA within the mouse oocyte. We achieved high rates of mutagenesis of the mouse Zic2 gene in all backgrounds examined including outbred CD1 and inbred C3H and C57BL/6J. Founder mutant Zic2 mice (eight independent alleles, with frameshift and deletion mutations) were created in C3H and C57BL/6J backgrounds. These mice transmitted the mutant alleles to the progeny with 100% efficiency, allowing the creation of inbred lines. Mutant mice display a curly tail phenotype consistent with Zic2 loss-of-function. The efficiency of site-specific germline mutation in the mouse confirm TALEN mediated mutagenesis in the oocyte to be a viable alternative to conventional gene targeting in embryonic stem cells where simple loss-of-function alleles are required. This technology enables allelic series of mutations to be generated quickly and efficiently in diverse genetic backgrounds and will be a valuable approach to rapidly create mutations in mice already bearing one or more mutant alleles at other genetic loci without the need for lengthy backcrossing.
Slow, Ultraslow, Stored, and Frozen Light (2002)

Get PDF Email Share Share with Facebook Tweet This Post on reddit Share with LinkedIn Add to CiteULike Add to Mendeley Add to BibSonomy Get Citation Copy Citation Text Yuri Rostovtsev, Olga Kocharovskaya, George R. Welch, and Marlan O. Scully, "Slow, Ultraslow, Stored, and Frozen Light," Optics & Photonics News 13(6), 44-48 (2002) Export Citation BibTex Endnote (RIS) HTML Plain Text Citation alert Save article
Maori Psychology: A Long Way from Imago, He Ara Roa Tonu (1) (2005)

Coassemblies with tailored functions, such as drug loading, tissue targeting and releasing, therapeutic and/or imaging purposes, and so on, have been widely studied and applied in biomedicine. <i>De novo</i> design of these coassemblies hinges on an integrated approach involving synergy between various design strategies, ranging from structure screening of combinations of "phthalocyanine-chemotherapeutic drug" molecules for molecular scaffolds, exploration of related fabrication principles to verification of intended activity of specific designs. Here, we propose an integrated approach combining computation and experiments to design from scratch coassembled nanoparticles. This nanocoassembly, termed NanoPC here, consists of phthalocyanine-based scaffolds hosting chemotherapeutic drugs, aimed at hypersensitive chemotherapy guided by photoimaging for targeting tumors. Our design starts from the selection of phthalocyanine derivatives that are not aggregation-prone, followed by computational screening of coassembled molecules covering various categories of chemotherapy drugs. To facilitate an efficient and accurate assessment of coassembly capabilities, we utilize small systems as surrogates to enable free-energy calculations at all-atom levels facilitated with enhanced sampling and statistical mechanics for efficient and accurate evaluation of coassembly ability. The final top NanoPC candidate, comprised of phthalocyanine PcL and cytarabine (CYT), can greatly increase the fluorescence intensity ratio of tumor/liver by 21.5 times and achieve higher antitumor efficiency in a pH-dependent manner. Therefore, the designing approach proposed here has a potential pattern, which can provide ideas and references for the design and development of coassembled nanodrugs with tailored functions and applications in biomedicine.
Potential radiosensitizing agents. 2. Synthesis and biological activity of derivatives of dinitroimidazole with oxiranes (1981)

A series of 1-substituted 2,4-dinitroimidazole analogues have been synthesized and tested for their radiosensitizing ability for selectively sensitizing hypoxic mammalian cells to the lethal effect of radiation. The reaction of 2,4-(5)-dinitroimidazole (1) with a variety of oxiranes upon heating in absolute ethanol yielded the expected 1-substituted 2,4-dinitroimidazoles (2) and also resulted in the formation of a novel class of isomeric nitroimidazo[2,1-b]oxazoles 3 and 4) by intramolecular cyclization. The results of radiosensitizing activity of these agents against hypoxic Chinese hamster cells (V-79) indicated that 2,4-dinitroimidazoles were better sensitizers than the nitroimidazo[2,1-b]oxazoles, suggesting the necessity of the 2-nitro function in the molecule. The 1-(2-hydroxy-3-methoxypropyl)-2,4-dinitroimidazole (2d) was found to be the most effective radiosensitizer of this series.
Human estrogen sulfotransferase (SULT1E1) pharmacogenomics: gene resequencing and functional genomics (2003)

1. Estrogens are used as drugs and estrogen exposure is a risk factor for hormone-dependent diseases such as breast cancer. Sulfate conjugation is an important pathway for estrogen metabolism. The sulfotransferase (SULT) enzyme SULT1E1 has the lowest K(m) values for estrogens and catecholestrogens of the 10 known human SULT isoforms. 2. We previously cloned and characterized the human SULT1E1 cDNA and gene as steps toward pharmacogenetic studies. In the present experiments, we set out to determine whether common, functionally significant genetic polymorphisms might exist for SULT1E1. As a first step, we 'resequenced' the eight SULT1E1 exons and exon-intron splice junctions as well as portions of the 5'-flanking region using DNA from 60 African-American and 60 Caucasian-American subjects. 3. In all, 23 polymorphisms, 22 single nucleotide polymorphisms (SNPs) and one insertion deletion were observed. There were three nonsynonymous coding SNPs (cSNPs) that altered the following encoded amino acids: Asp22Tyr, Ala32Val and Pro253His. Among these, 12 pairs of SNPs were tightly linked. In addition, 12 unambiguous SULT1E1 haplotypes were identified, including six that were common to both populations studied. 4. Transient expression in COS-1 cells of constructs containing the three nonsynonymous cSNPs showed significant decreases in SULT1E1 activity for the Tyr22 and Val32 allozymes, with corresponding decreases in levels of immunoreactive protein. There were no changes in levels of either activity or immunoreactive protein for the His253 allozyme. Apparent K(m) values of the Val32 allozyme for the two cosubstrates for the reaction, 17beta-estradiol and 3'-phosphoadenosine 5'-phosphosulfate, were not significantly different from those of the wild-type enzyme, but there was a two- to three-fold increase in K(m) values for the His253 allozyme and a greater than five-fold increase for the Tyr22 allozyme. 5. These observations raise the possibility that genetically determined variation in SULT1E1-catalyzed estrogen sulfation might contribute to the pathophysiology of estrogen-dependent diseases as well as variation in the biotransformation of exogenously administered estrogens.
Loss of sphingosine kinase‐1 activates the intrinsic pathway of programmed cell death: modulation of sphingolipid levels and the induction of apoptosis (2005)

Activation of sphingosine kinase-1 (SK1) by overexpression or agonist stimulation promotes cell proliferation, survival, and anti-apoptosis. Studies on the function of endogenous SK1 are lacking. Endogenous SK1 has been shown to be down-regulated under stress, and knockdown of the enzyme reduces the percentage of viable MCF-7 breast cancer cells (Taha, T. A. et al. 2004. J. Biol. Chem. 279, 20546-20554). In this study, we examined the mechanisms by which SK1 loss affects the growth of cells. Knockdown of the enzyme by small interfering RNA caused cell cycle arrest and induced apoptosis. Cell death involved effector caspase activation, cytochrome c release and Bax oligomerization in the mitochondrial membrane, thus placing SK1 knockdown upstream of the mitochondrial pathway of apoptosis. SK1 knockdown also induced significant increases in ceramide levels in whole cells and in mitochondria enriched fractions of cells. Inhibition of de novo sphingolipid biosynthesis with myriocin significantly attenuated Bax oligomerization and downstream caspase activation after SK1 loss. These studies for the first time implicate endogenous SK1 as an important survival enzyme in MCF-7 cells and link the biological consequences of knocking down the enzyme to its biochemical role as a regulator of sphingolipid metabolism.
Effect of charge on the interaction of site-specific photoimmunoconjugates with human ovarian cancer cells. (1996)

A marked effect of charge modification on the uptake and phototoxicity of a photoimmunoconjugate (PIC) was demonstrated. A site-specific conjugation strategy was developed to attach the photosensitizer chlorin(e6) (c(e6)) to the F(ab')2 fragment of the murine antiovarian cancer monoclonal antibody OC125. Poly-L-lysine linkers carrying c(e6) with a cationic charge or by polysuccinylation with an anionic charge were used and covalently attached to partially reduced antibody via a heterobifunctional reagent. PICs were purified by column chromatography and were also radiolabeled with 125I. PIC binding and uptake were studied with a human ovarian cancer cell line, NIH-OVCAR-5, and a nonantigen-expressing colon cancer cell line, SW1116, and the data were compared with the binding and uptake of nonspecific rabbit IgG PICs. PICs with both cationic and anionic charges preserved antigen binding as shown by competition studies with native antibody, but the cationic PIC had up to 17 times higher cellular uptake of c(e6), probably due to enhanced internalization. The ratio of c(e6) to 125I retained by the cells varied with the likelihood of internalization and lysosomal degradation. The phototoxicity of the PICs generally varied with their uptake, but a correlation was found between lysosomal hydrolysis as measured by an increased cellular ratio of c(e6):125I and increased relative phototoxicity. These data suggest cationic PICs may have advantages for photoimmunotherapy of disseminated intracavity cancer following local administration.
Photoacoustic flow measurements by use of laser-induced shape transitions of gold nanorods (2005)

A quantitative technique for flow measurements based on a wash-in analysis is proposed. The technique makes use of the shape dependence of the optical absorption of gold nanorods and the transitions in their shape induced by pulsed laser irradiation. The photon-induced shape transition of gold nanorods involves mainly a rod-to-sphere conversion and a shift in the peak optical absorption wavelength. The application of a series of laser pulses will induce shape changes in gold nanorods as they flow through a region of interest, with quantitative flow information being derived from the photoacoustic signals from the irradiated gold nanorods measured as a function of time. To demonstrate the feasibility of the technique, a Nd:YAG laser operating at 1064 nm was used for irradiation and a 1 MHz ultrasonic transducer was used for acoustic detection. The flow velocity ranged from 0.35 to 2.83 mm/s. Excellent agreement between the measured velocities and the actual velocities was demonstrated, with a linear regression correlation coefficient of 0.93. This study is a pioneer work on wash-in flow estimation in photoacoustic imaging.
The binding conformation of Taxol in β-tubulin: A model based on electron crystallographic density (2001)

The chemotherapeutic drug Taxol is known to interact within a specific site on beta-tubulin. Although the general location of the site has been defined by photoaffinity labeling and electron crystallography, the original data were insufficient to make an absolute determination of the bound conformation. We have now correlated the crystallographic density with analysis of Taxol conformations and have found the unique solution to be a T-shaped Taxol structure. This T-shaped or butterfly structure is optimized within the beta-tubulin site and exhibits functional similarity to a portion of the B9-B10 loop in the alpha-tubulin subunit. The model provides structural rationalization for a sizeable body of Taxol structure-activity relationship data, including binding affinity, photoaffinity labeling, and acquired mutation in human cancer cells.
Stability of Patch Methylation and Its Impact in Regions of Transcriptional Initiation and Elongation (1997)

CpG DNA methylation has previously been correlated with the suppression of transcription. The mechanism of this suppression is not understood, and many aspects of the temporal and positional relationships between the region of methylation and transcription have not yet been defined. Here, 12-kb stable replicating episomes that can be maintained in human somatic cells for weeks to months were used. Such a system allows more direct manipulation and is free from the positional effects attendant with the analysis of endogenous loci or integrated transgenes. By using these circular minichromosomes, patches of CpG methylation were created to include or exclude the regions of transcriptional initiation and elongation. I found that a 0.5-kb patch of methylation that covered the promoter suppressed expression only 2-fold and that a 1.9-kb patch of methylation that covered the coding portion of the gene (but not the promoter) suppressed expression about 10-fold. In contrast, methylation of the entire minichromosome except for the promoter or the coding portion suppressed transcription about 50- to 200-fold. I infer the following. Methylation of the 0.5-kb promoter fragment does not significantly affect transcription at the level of transcription factor binding or local chromatin structure. The dominant effect on transcription occurs when the length of methylated DNA is long, with little disproportionate effect of methylation of specific regions, such as that of initiation or elongation. I also found that the boundaries between these methylated and unmethylated regions remained stable for the many weeks that I monitored them.
Roles of intracellular cAMP and protein kinase A in the actions of dopamine and neurotensin on midbrain dopamine neurons (1992)

The role of intracellular cAMP and protein kinase A in dopamine-induced inhibition of dopamine neurons and the attenuation of this inhibition by neurotensin were studied in rat midbrain slices. Spontaneous activity of dopamine cells was recorded extracellularly from both the ventral tegmental area and the substantia nigra. Perfusion of the slices with 8-bromo-cAMP and forskolin significantly attenuated dopamine-induced inhibition, but neither blocked the inhibition completely. Neither SQ22536, an inhibitor of adenylate cyclase, nor H8, an inhibitor of protein kinase A, mimicked the inhibitory effect of dopamine on dopamine neurons, although they potentiated dopamine's effect. These results indicate that dopamine-induced inhibition of dopamine neurons can be affected by intracellular cAMP levels, but is unlikely to be mediated solely by inhibition of adenylate cyclase. The similarities between the effects of neurotensin and those of 8-bromo-cAMP and forskolin suggest that intracellular cAMP may be involved in the actions of neurotensin. This suggestion is supported by our findings that isobutyl-methylxanthine (an inhibitor of phosphodiesterases) potentiated the effect of neurotensin and SQ22536 and H8 antagonized it. Phorbol-12,13-dibutyrate (an activator of protein kinase C) did not mimic the effect of neurotensin, and H7 (an inhibitor of protein kinase C) did not reduce the effect, suggesting that protein kinase C is unlikely to be involved in the modulatory effect of neurotensin.
Mite‐antigen avoidance can reduce bronchial epithelial shedding in allergic asthmatic children (1998)

Exposure to house dust mite antigen can induce airway epithelial shedding even in subjects with low eosinophil airway infiltration, thus supporting the idea that epithelial damage in asthmatics sensitized to Dermatophagoides may be due to a proteolytic activity of the mite major antigens.
Discovery and Preclinical Characterization of INCB018424, a Selective JAK2 Inhibitor for the Treatment of Myeloproliferative Disorders. (2007)

Abstract Activating mutations in Janus kinase 2 (JAK2) have recently been identified in the majority of Philadelphia chromosome negative (Ph-) myeloproliferative disorders (MPDs). Importantly, constitutive JAK2 activation is oncogenic and, in murine models, recapitulates much of the pathobiology observed in MPD patients, suggesting that JAK2 inhibition may be of therapeutic benefit. Here we describe the identification and preclinical characterization of INCB018424, a potent, selective, and orally bioavailable inhibitor of the JAK2 now in clinical trials. INCB018424 was identified through an extensive medicinal chemistry effort designed to optimize potency, selectivity, pharmaceutical and pharmacokinetic properties. INCB018424 inhibits JAK2 at &lt;1 nM and demonstrated &gt;500–fold selectivity against a broad sampling of the kinome. The potency and selectivity of INCB018424 translated to exceptional cellular activity where it inhibited the proliferation of FDCP cells and BaF/3 cells expressing JAK2V617F with an IC50 of 100–130 nM, but not the proliferation of cell lines expressing activating mutations in either BCR-Abl or cKit (IC50 &gt; 25 and 4 mM, respectively). The effect of INCB18424 on cell proliferation correlated well with reduced levels of phosphorylated JAK2 and STAT5 in the BaF/3 cell model, suggesting that the effect is mediated by pharmacological inhibition of JAK-STAT pathway. Interestingly, the activation of endogenous wild-type JAKs - by the addition of IL-3 - shifted the potency of INCB018424 in the BaF/3 model greater than five fold suggesting that cells expressing the mutated form of JAK2 may be more sensitive to INCB018424. Indeed, using cells harvested from patients with Jak2V617F-positive polycythemia vera (PV) in colony forming assays, we observed that INCB018424 inhibited the cytokine-independent formation of erythroid progenitor colonies (n=3) with an IC50 of 67nM while normal colony formation from healthy donors (n=3) was inhibited 50% at &gt; 400 nM. Further, INCB018424 inhibited proliferation of PV patient samples (n=3) following ex vivo expansion of erythroid progenitors in serum free media, with an IC50 of 60 nM, similar to that observed in semi-solid media. In a mouse model of MPD, where implantation of BaF/3 cells expressing JAK2V617F results in rapid organomegaly and reduced survival, oral administration of INCB018424 was well tolerated and markedly reduced the splenomegaly. Using this animal model, we also demonstrated that selective JAK inhibition eliminates neoplastic cells from the spleen, liver, and bone marrow normalizing the histology of affected organs and significantly prolongs survival. As such, potent and selective JAK inhibitors such as INCB018424 hold great promise for the treatment of MPDs and other disease states associated with elevated JAK activity - a concept currently being tested clinically.
LSP1 Is the Major Substrate for Mitogen-activated Protein Kinase-activated Protein Kinase 2 in Human Neutrophils (1997)

In intact cells, mitogen-activated protein kinase-activated protein (MAPKAP) kinase 2 is rapidly activated by various cytokines, stresses, and chemotactic factors. The small heat shock protein p27 has been shown to be a substrate for MAPKAP kinase 2. Recently, we identified a novel substrate, designated p60, for MAPKAP kinase 2 in human neutrophils (Zu, Y.-L., Ai, Y., Gilchrist, A., Labadia, M. E., Sha'afi, R. I., and Huang, C.-K. (1996) Blood 87, 5287-5296). To further understand the signaling pathway of MAPKAP kinase 2, we have purified p60 from a heat-treated neutrophil lysate by DEAE-cellulose chromatography and SDS-polyacrylamide gel electrophoresis. Microsequencing of five peptides derived from purified p60 indicates that p60 is lymphocyte-specific protein 1 (LSP1). Furthermore antibodies specific for human and mouse LSP1 react with human and mouse p60. The sequence of human LSP1 indicates two serine residues at positions 204 and 252 as potential phosphorylation sites. The amino acid sequences surrounding these two sites are in agreement with the consensus sequence (Xaa-Xaa-Hyd-Xaa-Arg-Xaa-Xaa-Ser-Xaa-Xaa) for phosphorylation by MAPKAP kinase 2. Both serine residues in human LSP1 and the corresponding conserved serine residues in mouse LSP1 are in the basic C-terminal F-actin binding domain. Various fusion proteins of wild type and truncated mouse LSP1 with glutathione S-transferase were tested for their capacity to be phosphorylated by MAPKAP kinase 2. The results indicate that LSP1 is a substrate for MAPKAP kinase 2 in vitro and that the phosphorylation sites are located in the basic C-terminal domain of LSP1. Because both the small heat shock proteins and LSP1 are F-actin binding proteins, these results suggest a role for MAPKAP kinase 2 in the regulation of cytoskeletal structure or function.
Digital radiography (1988)

A quantitative comparison of the digital techniques reviewed in section 4.1-4.7 is difficult, for two reasons. Firstly, various authors have used slightly different techniques for assessing aspects of imaging performance (e.g. a variety of test objects for, and definitions of, spatial resolution). Secondly, with all imaging systems there exists an inter-relationship between spatial resolution, image acquisition time, image noise and dose. Some authors have chosen to emphasise one feature at the expense of others. Arnold (1982), in an overview of digital radiographic technology at that time, also noted the lack of standardisation of measurement techniques and exposure conditions, but nevertheless attempted a quantitative comparison of some aspects of digital radiographic systems with screen-film radiography and CT. The continuing developments in the field since then make a brief quantitative intercomparison of dubious value. Nevertheless, a qualitative summary of point, line and area exposure techniques is given in table 2 which incorporates many of the comments made by Arnold et al (1986) in a similar summary of digital radiographic systems.