Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We recently described 17 anti-CRP mAb, seven to native- (or conformational) and 10 to neo- (or sequence-determined) epitopes, including several anti-neo-CRP mAb specific for CRP peptide 199-206. In the present study, four new anti-native- and four new anti-neo-CRP mAb were generated and characterized by ELISA reactivity with native and modified human and rabbit CRP, as well as binding to pronase fragments of human CRP in Western blots. Assays with 17 synthetic CRP peptides identified anti-neo-CRP mAb specific for peptides 1-16, 14-24 and 137-152, respectively. The anti-neo-CRP mAb were reacted with fragments obtained by digesting CRP with multiple additional enzymes, including Staphylococcal V8 protease, trypsin, elastase, plasmin, thrombin and alpha-chymotrypsin. Native CRP was remarkably resistant to enzymic digestion, particularly in the presence of calcium, but was readily cleavable upon denaturation. Twenty-three informative fragments served to further distinguish mAb reactivity with at least four additional neo-CRP epitopes, which presumptively included residues in the regions of amino acids 22-45, 41-61, 114-121 and 130-138, respectively. The eight epitopes identified corresponded well with predicted regions of CRP antigenicity. In addition, at least six distinct native or conformation-determined epitopes were delineated. Reactivity of the anti-neo-CRP mAb with fragments of CRP generated by PMN enzymes indicated that regions sensitive to cleavage by neutrophil enzymes are located at approximately 3, 10 and 16 kD from the amino terminus of the CRP subunit. We expect that the anti-CRP mAb described and mapped herein will be useful tools for the elucidation of CRP structure and function.
Mol Immunol 1992 May
PMID:Localization of sequence-determined neoepitopes and neutrophil digestion fragments of C-reactive protein utilizing monoclonal antibodies and synthetic peptides. 137 44

Chemotactic and mitogenic activities of granulosa cells in developing follicles were studied. Immature rats were subcutaneously injected with 20 IU of pregnant mare's serum gonadotrophin and killed at various intervals after injection. The ovaries were removed and granulosa cells were isolated and cultured in a serum-free medium supplemented with insulin, transferrin and hydrocortisone. Chemotactic and mitogenic activities in the conditioned medium were determined. Our results demonstrated that in addition to mitogenic activity, chemotactic activity was also expressed in the conditioned medium of granulosa cells. Both activities increased with the maturity of follicles. A gel filtration analysis revealed that there were two peaks showing both mitogenic and chemotactic activities with a molecular size smaller than 5000. These peaks had various sensitivities to heat and trypsin treatment. In addition, the active component of both peaks was organic solvent-extractable. A thin-layer chromatography analysis indicated that the lipid component was not prostaglandin, estradiol or hydrocortisone.
Mol Cell Endocrinol 1992 Mar
PMID:Chemotactic and mitogenic activities of granulosa cells in developing follicles. 137

Amino acids in the serine proteinase inhibitor eglin c important for its inhibitory specificity and activity have been investigated by site-directed mutagenesis. The specificity of eglin c could be changed from elastase to trypsin inhibition by the point mutation Leu45----Arg (L45R) in position P1 [nomenclature according to Schechter and Berger (1967) Biochem. Biophys. Res. Commun. 27, 157-162]. Model building studies based on the crystal structure of mutant L45R [Heinz et al. (1991) J. Mol. Biol. 217, 353-371] were used to rationalize this specificity change. Surprisingly, the double mutant L45R/D46S was found to be a substrate of trypsin and various other serine proteinases. Multidimensional NMR studies show that wild-type eglin c and the double mutant have virtually identical conformations. In the double mutant L45R/D46S, however, the N-H bond vector of the scissile peptide bond shows a much higher mobility, indicating that the internal rigidity of the binding loop is significantly weakened due to the loss or destabilization of the internal hydrogen bond of the P1' residue. Mutant T44P was constructed to examine the role of a proline in position P2, which is frequently found in serine proteinase inhibitors [Laskowski and Kato (1980) Annu. Rev. Biochem. 49, 593-626]. The mutant remains a potent elastase inhibitor but no longer inhibits subtilisin, which could be explained by model building. Both Arg51 and Arg53, located in the core of the molecule and participating in the hydrogen bonding network with residues in the binding loop to maintain rigidity around the scissile bond, were individually replaced with the shorter but equally charged amino acid lysine. Both mutants showed a decrease in their inhibitory potential. The crystal structure of mutant R53K revealed the loss of two hydrogen bonds between the core and the binding loop of the inhibitor, which are partially restored by a solvent molecule, leading to a decrease in inhibition of elastase by 2 orders of magnitude.
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PMID:Changing the inhibitory specificity and function of the proteinase inhibitor eglin c by site-directed mutagenesis: functional and structural investigation. 139 Jun 62

Protease nexin-II (PN-II) is a potent chymotrypsin inhibitor that forms SDS-stable inhibitory complexes with epidermal growth factor binding protein, the gamma-subunit of nerve growth factor, and trypsin, and represents the secreted form of the amyloid beta-protein precursor (APP) that contains the Kunitz-type protease inhibitor domain. To determine the expression of PN-II within the peripheral nervous system, human dorsal root ganglia were processed for immunocytochemistry using well-characterized monoclonal antibodies against PN-II and for in situ hybridization studies using 35S-RNA PN-II probes for both APP751 and APP770. Highly specific immunoperoxidase staining of PN-II was demonstrated within the cytoplasm of dorsal root ganglia neurons and their processes in cryostat (fresh frozen) and vibratome (paraformaldehyde-fixed) sections. In situ hybridization using an anti-sense 35S-RNA PN-II probe demonstrated the presence of intense neuronal labeling. Labeling was not observed when the corresponding sense 35S-RNA PN-II probe was used. Although the precise functional role of PN-II/APP is not clear, the accumulation of amyloid beta-protein within the neuropil appears to be one of the earliest events in the pathogenesis of Alzheimer's disease (AD). Thus knowledge of the cell populations expressing the PN-II/APP gene would certainly be helpful for studies of the molecular mechanisms leading to the morphological and functional changes of AD. The results of this study clearly establish the expression of PN-II and its mRNA within the dorsal root ganglia neurons and their processes, and provide another point of departure for studies of the molecular mechanisms underlying the deposition of amyloid beta-protein and its relationships to the formation of neuritic plaques and neurofibrillary tangles.
Mol Chem Neuropathol 1992 Jun
PMID:Expression of protease nexin-II in human dorsal root ganglia. A correlative immunocytochemical and in situ hybridization study. 141 19

The distribution of O-linked oligosaccharides on the M(r) 55,000 glycoproteins, ZP3 alpha and ZP3 beta, of the porcine oocyte zona pellucida was examined. Purified preparations of endo-beta-galactosidase digested ZP3 alpha and ZP3 beta were reduced and carboxamidomethylated and digested with trypsin. When the trypsin digests were mapped by HPLC, each glycoprotein yielded only one N-acetylgalactosamine containing glycopeptide. Purification of the O-glycopeptides was achieved by a two-step protocol. Tryptic digests were applied to jacalin-agarose and specifically-bound O-glycopeptides (alpha OGP and beta OGP) were eluted with buffer containing 50 mM alpha-methylgalactoside as the haptenic sugar. Further purification of each O-glycopeptide was accomplished by reverse phase HPLC. Purified O-glycopeptides were characterized with respect to amino acid and carbohydrate compositions and sequenced by automated Edman degradation; alpha OGP was a 41-residue glycopeptide with three O-linked sugar chains. Sequence comparisons revealed a 75% identity between alpha OGP and a corresponding segment of rabbit rec55 zona protein; beta OGP was a 25-residue glycopeptide characterized by the presence of one N-linked and five O-linked sugar chains and a trypsin-resistant internal arginine residue. Sequence alignments revealed an 80% or greater identity between beta OGP and internal peptides of mouse, hamster and human ZP3 zona proteins. These studies demonstrate that in the case of ZP3 alpha and ZP3 beta, the pig homologues of rabbit rec55 and mouse ZP3, respectively, O-linked oligosaccharides are confined within delimited domains rather than widely dispersed on the polypeptide backbone. Such clustering of O-linked oligosaccharides may represent an essential determinant of the structure and biological activity of zona proteins.
Mol Reprod Dev 1992 Oct
PMID:Porcine oocyte zona pellucida M(r) 55,000 glycoproteins: identification of O-glycosylated domains. 141 87

A 14.5 kDa barley endosperm protein that is a major allergen in baker's asthma disease, as previously shown by both in vitro (IgE binding) and in vivo tests, has been identified as a glycosylated monomeric member of the multigene family of inhibitors of alpha-amylase/trypsin from cereals. A cDNA encoding this allergen (renamed BMAI-1) has been isolated and characterized. The deduced sequence for the mature protein, which is 132 residues long, is identical in its N-terminal end to the 20 amino acid partial sequence previously determined from the purified allergen, and fully confirms that it is a member of the multigene family of cereal inhibitors. Southern-blot analysis of wheat/barley addition lines using the insert in the BMAI-1 cDNA clone as a probe, has led to the location of the allergen gene (Iam1) in barley chromosome 2, while another related member of this protein family, the barley dimeric alpha-amylase inhibitor BDAI-1 gene (Iad1) has been located in chromosome 6. Iam1 is the first member of this inhibitor family in cereals to be assigned to chromosome group 2, thus extending the dispersion of genes in the family to five out of the seven homology groups of chromosomes in wheat and barley (chromosomes 2, 3, 4, 6 and 7).
Plant Mol Biol 1992 Nov
PMID:A major barley allergen associated with baker's asthma disease is a glycosylated monomeric inhibitor of insect alpha-amylase: cDNA cloning and chromosomal location of the gene. 142 Nov 48

The adult newt cardiac ventricular myocyte has been successfully placed in cell culture and has been shown to undergo in vitro DNA synthesis. Although several growth factors have been reported to increase DNA synthesis in cardiac myocytes in vitro, PDGF has not been reported to do so, but has been shown to be active in other systems. Ventricles were removed from the adult red-spotted newt and were enzymatically and mechanically dissociated in a solution containing trypsin and collagenase. Cells were preplated on to plastic to remove non-myocytes. Myocytes were then plated onto laminin. Groups of myocytes were fed control medium and medium containing porcine PDGF. Myocytes were given 1 microCi/ml of tritiated thymidine 6 or 24 h before fixation. Control myocytes showed a peak DNA synthesis at 12-14 days in culture. One ng/ml of PDGF increased DNA synthesis significantly to 22% above control. Myocytes responded to PDGF with significantly increased DNA synthesis in about 12 h. PDGF did not induce earlier DNA synthesis, but increased synthesis at all days of culture tested. These results indicate that PDGF acts upon cardiac myocytes, increasing their DNA synthesis.
J Mol Cell Cardiol 1992 Sep
PMID:Stimulation of DNA synthesis by PDGF in the newt cardiac myocyte. 143 20

At least 3 structural protein precursors of the eggshell are synthesized and stockpiled in the extensive vitelline cells of the liver fluke Fasciola hepatica L. One of these, vitelline protein B, consists of a closely related family of proteins that owes its apparent electrophoretic heterogeneity to variations in the Tyr to DOPA conversion as well as to subtle variations in the primary sequence. The efficiency of the Tyr to DOPA conversion ranges from a maximum of about 90% to a minimum of 55% in the protein. Trypsin digestion in borate buffer at pH 8 was used to produce DOPA-peptides for sequencing. Notably, trypsin does not cleave Arg/Lys-DOPA sequences at borate concentrations greater than 0.15 M. Peptides with DOPA-containing sequences most frequently have flanking amino acids such as Lys, Ser, or Asp on the N-terminal side and Gly or Asp on the C-terminal side. All protein variants fall within a narrow molecular weight range (30-33 kDa), a pI range of 6.9 to 8.3, and the collective majority would appear to share a common N-terminal sequence up to residue 28. The results suggest some combination of the following: variations in post-translational hydroxylation, alternative post-transcriptional splicing and/or the existence of multiple gene copies of eggshell precursors. The latter have been shown to occur in the blood fluke Schistosoma mansoni [15].
Mol Biochem Parasitol 1992 Sep
PMID:Eggshell precursor proteins of Fasciola hepatica, II. Microheterogeneity in vitelline protein B. 143 55

Chromosomal proteins HMG-14 and HMG-17 have a modular structure. Here we examine whether the putative nucleosome-binding domain in these proteins can function as an independent module. Mobility shift assays with recombinant HMG-17 indicate that synthetic molecules can be used to analyze the interaction of this protein with the nucleosome core. Peptides corresponding to various regions of the protein have been synthesized and their interaction with nucleosome cores analyzed by mobility shift, thermal denaturation and DNase I digestion. A 30 amino acid long peptide, corresponding to the putative nucleosome-binding domain of HMG-17, specifically shifts the mobility of cores as compared to free DNA, elevates the tm of both the premelt and main melt of the cores and protects from DNase I digestion the same nucleosomal DNA sites as the intact protein. The binding of both the peptide and the intact protein is lost upon digestion of the histone tails by trypsin. The nucleosomal binding sites of the peptide appear identical to those of the intact protein. Thus, a region of the protein can acts as an independent functional domain. This supports the notion that HMG-14 and HMG-17 are modular proteins. This finding is relevant to the understanding of the function and evolution of HMG-14/-17, the only nucleosome core particle binding proteins known to date.
J Mol Biol 1992 Nov 20
PMID:Nucleosome core binding region of chromosomal protein HMG-17 acts as an independent functional domain. 145 55

Peptides containing either one, two or three of the three zinc-finger motifs from the yeast transcription factor SWI5 have been prepared by expression in Escherichia coli. The DNA binding characteristics of these peptides were investigated, and a two-dimensional nuclear magnetic resonance (n.m.r.) study undertaken to establish the three-dimensional structures of the two-finger peptide. The peptide containing fingers 1 and 2 binds sequence specifically to two thirds of the DNA binding site recognized either by intact SWI5 or by the isolated three-finger peptide, and hence has the correct tertiary fold for DNA recognition. These results also establish the polarity of DNA binding, since the N-terminal two fingers of SWI5 bind to the 5' end of the DNA binding site. Mild proteolysis of the three-finger peptide using trypsin results in a small number of discrete products, which is consistent with the presence of three structured mini-domains. Nearly complete n.m.r. signal assignments were obtained for two peptides containing finger 2 alone or fingers 1 + 2. Comparison of two-dimensional spectra of these peptides and others clearly shows that the NOE enhancements and chemical shifts characteristic of each finger are quite insensitive to the presence or absence of neighbouring fingers. This clearly indicates that adjacent zinc-finger domains are structurally independent in these peptides from SWI5. However, there must be some steric limitations on the possible relative orientations of the fingers, and to establish limits for these a set of structures for the peptide containing fingers 1 + 2 was calculated using the YASAP simulated annealing protocol in conjunction with n.m.r.-based constraints. A more detailed description of the three-dimensional structures of finger 1 and finger 2, and their relationship to other previously determined structures of single zinc-fingers, is given in the accompanying paper.
J Mol Biol 1992 Nov 20
PMID:Adjacent zinc-finger motifs in multiple zinc-finger peptides from SWI5 form structurally independent, flexibly linked domains. 145 67


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