bluuebunny/crossref_metadata_2025_split huggingface dataset:
A camel milk whey protein rich in half‐cystine
Authors: BEG Obaid Ullah , von BAHR‐LINDSTRÖM Hedvig , ZAIDI Zafar H. , JÖRNVALL Hans 
Published: March 2005

The amino acid sequence of a recently isolated camel milk protein rich in half‐cystine has been determined by peptide analyses. The 117‐residue protein has 16 half‐cystine residues, concluded to correspond to disulfide bridges and suggesting a tight conformation of the molecule.Comparisons of the structure with those of other proteins reveal several interesting relationships. The camel protein is clearly homologous with a previously reported rat whey phosphoprotein of possible importance for mammary gland growth regulation, and with a mouse protein of probable relationship to neurophysins. The camel, rat and mouse proteins may represent species variants from a rapidly evolving gene. Residue identities in pairwise comparisons are 40% for the camel/rat proteins and 33% for the camel/mouse proteins, with 38 positions conserved in all three forms. The camel protein also reveals an internal repeat pattern similar to that for the other two proteins.The homology between the three milk whey proteins has wide implications for further relationship. Thus, previously noticed similarities, involving either of the milk proteins, include limited similarities to casein phosphorylation sites for the camel protein, to neurophysins in repeat and half‐cystine patterns for the mouse and rat proteins, and to an antiprotease for the rat protein. These similarities are reinforced by the camel protein structure and the recognition of the three whey proteins as related. Finally a few superficial similarities with the insulin family of peptides and with some other peptides of biological importance are noticed. Combined, the results relate the camel protein in a family of whey proteins, and extend suggestions of relationships with some binding proteins.
Differential inhibition of prostaglandin and superoxide production by dexamethasone in primary cultures of rat Kupffer cells
Authors: DIETER Peter , SCHULZE‐SPECKING Agnes , DECKER Karl 
Published: March 2005

Dexamethasone inhibited the stimulus‐induced prostaglandin E2 formation by rat Kupffer cells in primary culture, e.g. after treatment with zymosan, phorbol ester, calcium ionophore A23187, platelet‐activating factor or lipopolysaccharide. Prostaglandin E2 production from added free arachidonic acid was not influenced by the hormone. The time course, as well as the partial inhibition of the hormone effect by actinomycin D and cycloheximide, point to the hormone‐induced formation of a protein which regulates phospholipase A2.The hormone did not affect the phagocytotic activity of the Kupffer cells. The quantity of [3H]arachidonic acid incorporated into phospholipids was also not altered by dexamethasone. After stimulation with zymosan, [3H]arachidonic acid was liberated from phosphatidylcholine only.Superoxide generation by rat Kupffer cells was induced by zymosan, phorbol ester and, to a much smaller extent, by platelet‐activating factor. A23187 and lipopolysaccharide were without effect. In contrast to prostaglandin formation, the generation of superoxide was not influenced by dexamethasone.These results indicate that in cultured rat Kupffer cells prostaglandin formation and superoxide generation are independently triggered processes.
Modification of the actin interface of skeletal myosin subfragment‐1 by treatment with dibromobimane
Authors: MORNET Dominique , UE Kathleen , CHAUSSEPIED Patrick , MORALES Manuel F. 
Published: March 2005

Recently, by treating the head portion of skeletal myosin subfragment‐1 (S1) with the bifunctional agent dibromobimane, we introduced an intramolecular covalent cross‐link which resulted in the stabilisation of an internal loop in the heavy chain structure of the head [Mornet et al. (1984) Proc. Natl Acad. Sci. USA 82, 1658–1662]. In order to define the functional properties of this new S1 conformational state, we have first determined the experimental conditions for the optimum modification of S1 by dibromobimane. We finally settled on a 60% yield of cross‐linked S1.Because the modification occurs between the 50‐kDa and the 20‐kDa tryptic heavy chain fragments which have been postulated to be involved in the interaction of native S1 with actin, we have investigated the association of dibromobimane‐treated S1 with actin, using chemical cross‐linking of their rigor complex with 1‐ethyl‐3‐[3‐(dimethylamino)propyl] carbodiimide. The cross‐linked species obtained were analyzed by polyacrylamide gel electrophoresis and compared with those known for unmodified S1. The carbodiimide‐catalyzed linkage between actin and dibromobimane‐modified S1 led to a singlet protein band migrating with an apparent molecular mass of 155 kDa, in contrast to the usual doublet bands of 175 kDa and 185 kDa produced with native S1. This result suggests that a change has occurred at the actin interface on the dibromobimane‐treated S1 heavy chain.The covalent complex generated by carbodiimide cross‐linking between actin and dibromobimane‐modified S1 (27‐kDa + 50‐kDa + 20‐kDa fragments) was submitted to chemical hydrolysis with hydroxylamine. The nature of the products identified is consistent with the conclusion that the internal freezing of the heavy chain structure by dibromobimane induces the loss of the ability to cross‐linkage of the actin site on the 20‐kDa domain but does not affect the conformation of the second site on the 50‐kDa segment, which becomes the unique actin region cross‐linkable by actin.
The aromatic 1H‐NMR spectrum of plasminogen kringle 4
Authors: RAMESH Vasudevan , GYENES Marianne , PATTHY László , LLINÁS Miguel 
Published: March 2005

The isolated kringle 4 domain of human plasminogen has been compared with homologous structures from bovine and porcine sources, both free and in the presence of the ligand 6‐aminohexanoic acid, by two‐dimensional 1H‐NMR spectroscopies at 300 MHz and 600 MHz. The chemical‐shift‐correlated, spin‐echo‐correlated, and double‐quantum‐correlated aromatic spectra of the three proteins reveal that the globular conformation of the fourth kringle is closely maintained throughout the set of homologs. Direct comparison shows that the three conserved Trp residues (at sites 25, 62 and 72) which exhibit highly non‐degenerate subspectra, find themselves in similar intramolecular environments. In particular, proton Overhauser experiments reveal that the close steric interaction between the Trp‐II (Trp62 or Trp25) indole group and the aromatic ring at site 74 (Tyr74 or Phe74) is strictly preserved. This feature forces the kringle inner loop, closed by the Cys51‐Cys75 link, to fold back onto itself so as to place the site 74 residue proximal to the Cys22‐Cys63 bridge.Single‐residue substitutions enable unambiguous assignments of His‐I to His3, Tyr‐III to Tyr41 and Tyr‐IV to Tyr74. From this direct evidence, comparison with the kringle 1 spectrum, and the previously reported chemical modification of Tyr‐II (Tyr50) [Trexler M., Bänyai L., Patthy L., Pluck N. D. & Williams R. J. P. (1985) Eur. J. Biochem. 152, 439–446], Tyr‐I and Tyr‐V (the latter, an immobile ring on the 600‐MHz time scale) could be assigned to Tyr2 and Tyr9, respectively. Since Trp‐III has previously been assigned to Trp72 at the lysine‐binding site, the present study completes the assignment of 10 out of 12 aromatic spin systems in the kringle 4 1H‐NMR spectrum; the only ambiguity which remains concerns the Trp‐I and Trp‐II indole spin systems, which are totally identified but as yet only tentatively assigned to Trp25 and Trp62, respectively.
Structural analysis of the sequence coding for an inducible 26‐kDa protein in human fibroblasts
Authors: HAEGEMAN Guy , CONTENT Jean , VOLCKAERT Guido , DERYNCK Rik , TAVERNIER Jan , FIERS Walter 
Published: March 2005

When human fibroblast cells were stimulated with poly(I) · poly(C) in the presence of cycloheximide for the production of interferon‐β (IFN‐β), a 26‐kDa protein could be immunoprecipitated by antiserum raised against partially purified human IFN‐β [Content, J., De Wit, L., Pierard, D., Derynck, R., De Clercq, E. & Fiers, W. (1982) Proc. Natl Acad. Sci. USA 79, 2768–2772]. In our hands this 26‐kDa protein showed no antiviral activity. Other investigators have, however, reported the presence in the same conditions of a second type of IFN, a socalled β2 species [Weissenbach, J., Chernajovsky, Y., Zeevi, M., Shulman, L., Soreq, H., Nir, U., Wallach, D., Perricaudet, M., Tiollais, P. & Revel, M. (1980) Proc. Natl Acad. Sci. USA 77, 7152–7156] of which the mRNA structure and protein characteristics strongly suggests identity with the 26‐kDa product.In this paper we describe the nucleotide sequence of the 26‐kDa cDNA and part of the corresponding genomic clone. The cDNA clones were isolated from a library made with mRNA from induced human fibroblasts. As, however, the information thus obtained was still incomplete, genomic clones were isolated from a total human DNA library. In this way, the entire region coding for the 26‐kDa protein was established, as well as the neighbouring sequences including the inducible promoter area. From the deduced polypeptide sequence a number of characteristics of the 26‐kDa protein can be explained. It turns out that the 26‐kDa protein gene and the socalled ‘IFN‐β2’ gene are identical. However, extensive homology searches indicate that the 26‐kDa protein does not show statistically significant sequence homology with any known interferon species. Hence, the question of whether the 26‐kDa product represents a novel IFN species remains open.
3‐Azi‐1‐methoxybutyl D‐maltooligosaccharides specifically bind to the maltose/maltooligosaccharide‐binding protein of Escherichia coli and can be used as photoaffinity labels
Authors: THIEME Roland , LAY Helga , OSER Andreas , LEHMANN Jochen , WRISSENBERG Sabine , BOOS Winfried 
Published: March 2005

Maltooligosaccharides with two to six (α1–4)‐linked glucose residues, carrying at their reducing end a 3‐azi‐1‐methoxybutyl group in either α or in β glycosidic linkage, were synthesized. These maltooligosaccharide analogues inhibit maltose uptake via the maltose‐binding‐protein‐dependent transport system in Escherichia coli. The concentration of half‐maximal inhibition of maltose transport, at 15 nM concentration, decreases with increasing chain length of the analogue, levelling off at 40 μM after a chain length of four glucose residues in the α series and at 350 μM after a chain length of three glucose residues in the β series. The inhibition of maltose transport occurs at the level of the periplasmic maltose‐binding protein. 3‐Azi‐1‐methoxybutyl α‐D‐[3H]maltotrioside was bound by the maltose‐binding protein with a Kd of 0.18 mM. Irradiation at 350 nm of purified maltose‐binding protein in the presence of 4 μM of this substrate labeled the protein covalently; labeling was prevented by 1 mM maltose. Using a crude preparation of periplasmic proteins two proteins were labeled, the maltose‐binding protein and α‐amylase.Thus, 3‐azi‐1‐methoxybutyl α‐D‐maltooligosaccharides are potent photoaffinity labels for proteins with maltooligosaccharides‐binding sites.
Porcine submaxillary mucin contains α2→3‐ and α2→6‐linked N‐acetyl‐ and N‐glycolyl‐neuraminic acid
Authors: SAVAGE Angela V. , KOPPEN Piet L. , SCHIPHORST Wietske E. C. M. , TRIPPELVITZ Leo A. W. , VAN HALBEEK Herman , VLIEGENTHART Johannes F. G. , VAN DEN EIJNDEN Dirk H. 
Published: March 2005

Four acidic trisaccharides have been obtained by alkaline borohydride reductive cleavage (β‐elimination) of a fraction of porcine submaxillary mucin precipitating at 60 – 75% ethanol. Their structures have been investigated using the techniques of methylation analysis involving gas‐liquid chromatography/mass spectrometry along with high‐resolution 1H‐NMR analysis. Two of the four oligosaccharides, B1 – B4, contain N‐acetylneuraminic acid (NeuAc) while two contain N‐glycolylneuraminic acid (NeuGc). The following structures are proposed for the acidic trisaccharide fraction: (B1) NeuAcα(2→3)Galβ(1→3)GalNAcol, (B2) NeuGcα(2→3)Galβ(1→3)GalNAcol, (B3)Galβ(1→3)[NeuAcα(2→6)]GalNAcol and (B4) Galβ(1→3)[NeuGcα(2→6)]GalNAcol, (GalNAcol = reduced N‐acetylgalactosamine.These oligosaccharides were present in a molar ratio of 69:22:4:5. Although oligosaccharide B4 has previously been found in porcine submaxillary mucin B1, B2 and B3 have not. Furthermore, oligosaccharide B2 is a novel structure.
ATP synthase complex from beef heart mitochondria
Authors: FREISLEBEN Hans‐Joachim , ZIMMER Guido 
Published: March 2005

ATPase/ATP synthase preparations originally contain protein bands in the 28–31‐kDa region. The present study demonstrates separation of the band at 29 kDa (adenine nucleotide translocator) from a band at approximately 31 kDa. In cholate/ammonium sulfate‐extracted ATP synthases removal of the 31‐kDa band results in decrease of ATP–Pi exchange and oligomycin sensitivity of the ATPase activity.It is suggested that the protein band at 31 kDa is heterogeneous, containing diverse activities, the identities of which are yet to be determined.
The Na+/H+ exchange system in glial cell lines
Authors: JEAN Thierry , FRELIN Christian , VIGNE Paul , LAZDUNSKI Michel 
Published: March 2005

1. The properties of the Na+/H+ exchange system in the glial cell lines C6 and NN were studied from 22Na+ uptake experiments and measurements of the internal pH (pHi) using intracellularly trapped biscarboxyethylcarboxyfluorescein. In both cell types, the Na+/H+ exchanger is the major mechanism by which cells recover their pHi after an intracellular acidification. The exchanger is inhibited by amiloride and its derivatives. The pharmacological profile (ethylisopropylamiloride > amiloride > benzamil) is identical for the two cell lines. Both Na+ and Li+ can be exchanged for H+. Increasing the external pH increases the activity of the exchanger in the two cell lines. In NN cells the external pH dependence of the exchanger is independent of the pHi. In contrast, in C6 cells, changing the pHi value from 7.0 to 6.5 produces a pH shift of 0.6 pH units in the external pH dependence of the exchanger in the acidic range. Decreasing pHi activates the Na+/H+ exchanger in both cell lines.2. Increasing the osmolarity of the external medium with mannitol produces an activation of the exchanger in C6 cells, which leads to a cell alkalinization. Mannitol action on 22Na+ uptake and the pHi were not observed in the presence of amiloride derivatives. Mannitol produces a modification of the properties of interaction of the antiport with both internal and external H+. It shifts the pHi dependence of the system to the alkaline range and the external pH (pHo) dependence to the acidic range. It also suppresses the interdependence of pHi and pHo controls of the exchanger's activity. NN cells that possess an Na+/H+ exchange system with different properties do not respond to mannitol by an increased activity of the Na+/H+ exchanger. The action of mannitol on C6 cells is unlikely to be mediated by an activation of protein kinase C.
Association of messenger RNA with the cytoskeletal framework in rat L6 myogenic cells
Authors: PRAMANIK Saroj K. , WALSH Richard W. , BAG Jnanankur 
Published: March 2005

The distribution of mRNA between the detergent‐soluble and insoluble (cytoskeleton) fractions in rat L6 myoblast and myotube cells was examined. Approximately 85% of cytoplasmic mRNA in both myoblasts and myotubes was found associated with the cytoskeletal framework. The cytoskeleton‐bound mRNA was present as polysomes. In contrast, the mRNA of the detergent‐soluble fraction was not associated with ribosomes and was thus considered to be the repressed population. The association of mRNA with the cytoskeletal framework was not affected by treatments leading to dissociation of polysomes. Differential distribution of mRNA between the soluble and cytoskeleton‐bound fractions was analyzed by in vitro translation. The mRNAs coding for polypeptides of molecular masses 40 kDa and 60 kDa were preferentially enriched in the soluble fraction.The nature of binding between mRNA and the cytoskeletal framework was examined following in vivo cross‐linking of RNA and protein by irradiating muscle cells with ultraviolet light. It was observed that this treatment covalently linked RNA and the neighbouring protein moieties without any detectable damage to the cytoskeletal framework, as measured by the distribution of RNAs and proteins between the cytoskeleton and soluble fractions. Analysis of the polypeptide moieties cross‐linked to the mRNA have shown that a large number of polypeptides of molecular masses between 15–220 kDa were associated with both cytoskeleton‐bound and soluble mRNAs. The polypeptide moieties of these mRNA‐protein complexes were not only similar in the cytoskeleton and soluble mRNA‐protein complexes but also were similar between myoblasts and myotubes. However, one polypeptide of 165 kDa was preferentially associated with the cytoskeleton‐bound mRNA‐protein complexes. Interestingly this 165‐kDa polypeptide was also preferentially enriched in the total proteins from the cytoskeleton fraction. This result suggests a possible role of the 165‐kDa polypeptide in association between mRNA and the cytoskeletal framework. To examine the mechanism of interaction between mRNA and the cytoskeletal framework we have reported here a ghost monolayer transcription system from myotubes. This transcription system was able to synthesize rRNA and mRNA. The mRNA transcribed in vitro was preferentially associated with the cytoskeleton structure present in the ghost monolayer system.
Nucleotide sequence and characteristics of the gene for l‐lactate dehydrogenase of Thermus caldophilus GK24 and the deduced amino‐acid sequence of the enzyme
Authors: KUNAI Kenji , MACHIDA Masayuki , MATSUZAWA Hiroshi , OHTA Takahisa 
Published: March 2005

The gene for L‐lactate dehydrogenase (LDH) (EC 1.1.1.27) of Thermus caldophilus GK24 was cloned in Escherichia coli using synthetic oligonucleotides as hybridization probes. The nucleotide sequence of the cloned DNA was determined. The primary structure of the LDH was deduced from the nucleotide sequence. The deduced amino acid sequence agreed with the NH2‐terminal and COOH‐terminal sequences previously reported and the determined amino acid sequences of the peptides obtained from trypsin‐digested T. caldophilus LDH. The LDH comprised 310 amino acid residues and its molecular mass was determined to be 32808. On alignment of the whole amino acid sequences, the T. caldophilus LDH showed about 40% identity with the Bacillus stearothermophilus, Lactobacillus casei and dogfish muscle LDHs. The T. caldophilus LDH gene was expressed with the E. coli lac promoter in E. coli, which resulted in the production of the thermophilic LDH. The gene for the T. caldophilus LDH showed more than 40% identity with those for the human and mouse muscle LDHs on alignment of the whole nucleotide sequences.The G + C content of the coding region for the T. caldophilus LDH was 74.1%, which was higher than that of the chromosomal DNA (67.2%). The G + C contents in the first, second and third positions of the codons used were 77.7%, 48.1% and 95.5% respectively. The high G + C content in the third base caused extremely non‐random codon usage in the LDH gene. About half (48.7%) the codons in the LDH gene started with G, and hence there were relatively high contents of Val, Ala, Glu and Gly in the LDH. The contents of Pro, Arg, Ala and Gly, which have high G + C contents in their codons, were also high. Rare codons with U or A as the third base were sometimes used to avoid the TCGA sequence, the recognition site for the restriction endonuclease, TaqI. Two TCGA sequences were found only in the sequence of CTCGAG (XhoI site) in the sequenced region of the T. caldophilus DNA. There were three segments with similar sequences in the two 5′ non‐coding regions, probably the promoter and ribosome‐binding regions, of the genes for the T. caldophilus LDH and the Thermus thermophilus 3‐isopropylmalate dehydrogenase.
Substitution of uridine in vivo by the intrinsic photoactivable probe 4‐thiouridine in Escherichia coli RNA
Authors: FAVRE Alain , BEZERRA Roberto , HAJNSDORF Eliane , DUBREUIL Yolande LEMAIGRE , EXPERT‐BEZANÇON Alain 
Published: March 2005

In vivo incorporation of the uridine‐photoactivable analogue, 4‐thiouridine, into the ribosomal RNA of an Escherichia coli pyrD strain has been demonstrated. It is highly dependent on the exogeneous uridine and 4‐thiouridine concentrations as well as on temperature. We have defined conditions allowing the substitution of 13 ± 2% of the uridine residues in bulk RNA by 4‐thiouridine. On a high‐Mg2+ sucrose gradient, 33 ± 3% of ribonucleic particles sediment as 70S ribosomes, the remaining being in the form of non‐associated 50S and 30S particles containing immature rRNA. The thiolated 70S ribosomes tolerate a 4–5% substitution level (40 thiouridine molecules/particle).Surprisingly, 3–4% of ribosomal proteins, about two protein molecules/particle, were spontaneously covalently bound to 4‐thiouridine‐substituted rRNA. Specific 366‐nm photoactivation increased this proportion to 10–12%, i.e. up to six or seven ribosomal protein molecules/particle. The photochemical cross‐linking proceeds with aparent first‐order kinetics with a quantum yield close to 5 × 10–3. Although extensive photodynamic breakage of rRNA occurs under aerobic conditions, both the kinetics and yield of ribosomal protein cross‐linking were independent of oxygenation conditions. The thiolated (4.5%) 70S ribosomes allowed the poly(U)‐directed poly(Phe)synthesis at 48% the control rate. Photoactivation decreased this activity to 28% and 10% when performed under nitrogen and in aerated conditions, respectively.
Regions necessary for pH‐dependent assembly of gizzard myosin rod
Authors: KUMON Akira , KUBA Masako , MURAKAMI Noriko , YASUDA Seiji , TAKASHIMA Toshinobu , MATSUMURA Sueo , SUEZAKI Yukio 
Published: March 2005

Aggregated and disaggregated forms of gizzard myosin rod and its fragments in various concentrations of NaCl (0–0.30 M) at various pH (7.4–8.6) were distinguished from each other by their permeability through a Sepharose 4B column.

The rod existed in three forms, namely: large aggregates impermeable to the column, small aggregates eluted at the void volume of the column and a disaggregated monomer which penetrated the column. The relative proportions of the three forms varied depending on the salt concentration and pH. The monomeric rod was detected in NaCl solutions above 0.20 M and its relative proportion at 0.25 M NaCl was larger than those of the small and large aggregates. The small aggregates of the rod were predominant at below 0.05 M NaCl and, upon decrease in pH from 8.6 to 7.4, these small aggregates in NaCl solutions between 0.10 M and 0.15 M were replaced by the large aggregates.
Light meromyosin, which corresponded to the C‐terminal two‐thirds of the rod, existed exclusively as large aggregates in NaCl solutions below 0.15 M; increase of NaCl concentration to above 0.20 M resulted in the formation of its monomer, instead of the large aggregates. In contrast to the rod, no small aggregated form of the light meromyosin was detected.
Truncated light meromyosin which had lost a small segment from either the C‐terminal or N‐terminal of light meromyosin was eluted only as a monomer in any NaCl concentration at any pH.
It may be deduced from the above results that a small segment in the light meromyosin is requisite for the assembly of both rod and light meromyosin in the NaCl solutions below 0.15 M and that the relative proportion of small and large aggregates of the rod is determined in a pH‐dependent manner by the subfragment 2 segment, the N‐terminal third of the rod.