Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.1 (chymotrypsin)
 10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Monoclonal antibodies were raised that specifically recognize the NH2-terminal neoepitope sequence present in link protein cleavage products derived from stromelysin-degraded proteoglycan aggregate. Competitive enzyme-linked immunosorbent assay, using synthetic peptides as inhibitors, showed that one of these antibodies (CH-3) required, for antibody recognition, the free NH2-terminal amino acid isoleucine (residue 17 of the intact protein) in the sequence NH2-IQAENG at the stromelysin cleavage site of link protein 3. Human proteoglycan aggregate was digested with recombinant human stromelysin, bovine chymotrypsin, bovine trypsin, and porcine elastase, and their respective link protein degradation products were tested for immunoreactivity with antibody CH-3. Only stromelysin- and chymotrypsin-generated link protein 3 were recognized by antibody CH-3. Both of these enzymes generate link protein NH2 termini with the sequence 17IQAENG. . .; hence these studies indicated that monoclonal antibody CH-3 recognized this neoepitope sequence in only specific proteolytically modified link protein molecules. Since the occurrence of link protein 3 increases with aging, the incidence of CH-3 epitope in proteoglycans isolated from human knee articular cartilage of individuals of different ages was investigated. The prevalence of CH-3 epitope was found to be highest in newborn and adolescent articular cartilage samples. However, little CH-3 epitope was detected in older adult cartilage, although considerably more link protein 3 was present in these samples. These results suggest that additional proteolytic agents are responsible for the increased occurrence of link protein degradation products with aging.
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PMID:Monoclonal antibodies recognizing protease-generated neoepitopes from cartilage proteoglycan degradation. Application to studies of human link protein cleavage by stromelysin. 137 86

Cartilage proteoglycan monomers associate with hyaluronic acid to form proteoglycan aggregates. Link protein, a glycoprotein interacting with both hyaluronic acid and proteoglycan, serves to stabilize the aggregate structure. The primary structure of the link protein has been determined with a view to defining its interaction with both hyaluronic acid and proteoglycan. Thus, the link protein has been digested with staphylococcal V8 protease, trypsin, and chymotrypsin and the resulting peptides characterized by amino acid composition and sequence. We have determined that the link protein is a single peptide with 339 amino acid residues. The protein core has a molecular weight of 38,564. There is one N-linked oligosaccharide at residue 41 with a molecular weight of approximately 2,500. There are five disulfide bonds which define three loops within the amino acid sequence. The loop nearest to the NH2-terminal contains 78 amino acids and is followed by a section of 42 amino acids between it and the second loop. The second and third loops display considerable homology with each other; they consist of 71 and 70 amino acids, respectively, each contain two disulfide bonds, and both loops possess, approximately centrally, an epitope for the species nonspecific anti-link protein monoclonal antibody, 8A4. These loops are separated by a short section of 27 amino acids. We speculate that these loops are functionally important in the interaction of link protein with hyaluronic acid, as they appear to be the most conserved regions of link protein between species.
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PMID:The primary structure of link protein from rat chondrosarcoma proteoglycan aggregate. 241 34

The ternary complex consisting of a 65-kDa peptide originating from the proteoglycan core protein and a 43-kDa link protein bound to hyaluronic acid was purified from a clostripain digest of the rat chondrosarcoma aggregating proteoglycan and 14C-carbamylated with potassium [14C]cyanate. At a pH of 8.0, 14C-carbamylation of the alpha-NH2 groups in the N-terminal amino acids was favored over carbamylation of epsilon-NH2 groups in the lysinyl residues for both the 65- and 43-kDa species. Two-dimensional tryptic peptide maps revealed a single major, distinctly different, fluorographic spot for each. These tryptic peptides had approximate masses of 4.5 kDa (from the 65-kDa species) and 3.0 kDa (from the 43-kDa species) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and each contained greater than 60% of the total radioactivity associated with its original polypeptide. Primary amino acid sequencing of the 65-kDa species gave a defined sequence for the first 4 N-terminal residues, whereas sequencing through the first 4 residues of a fully carbamylated species gave no dabsylated derivative for the first residue but identical residues in position 2-4 as for the noncarbamylated species and loss of radioactive derivative. Digests of 14C-carbamylated ternary complex with alpha-chymotrypsin resulted in a limit 14C-carbamylated 55-kDa species which contained greater than 85% of the radiolabel originally in the 65-kDa peptide. Similarly, trypsin generated two radiolabeled species, 60 and 58 kDa. These limit digest peptides (55, 60, 58 kDa) all contained the 4.5-kDa N-terminal tryptic peptide. Thus peptides removed from the 65-kDa peptide digestion with either alpha-chymotrypsin or trypsin were on the carboxyl end of the molecule.
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PMID:N-terminal carbamylation of the hyaluronic acid-binding region and the link protein from the chondrosarcoma proteoglycan aggregate. 353 3

Cartilage proteoglycan monomers associate with hyaluronic acid to form proteoglycan aggregates. Link protein, interacting with both hyaluronic acid and proteoglycan, serves to stabilize the aggregate structure. In the course of determining the primary structure of link protein, two peptides produced by digestion of rat chondrosarcoma link protein with trypsin or chymotrypsin have been selectively purified by immunoaffinity chromatography on a column of monoclonal anti-link protein antibody (8A4) immobilized to Sepharose 4B. These peptides have been sequenced using the double-coupling dimethylaminoazobenzene isothiocyanate/phenyl isothiocyanate procedure. A consensus sequence, Cys-X-Ala-Gly-Trp-Leu-X-Asp-Gly-Ser-Val-X-Tyr-Pro-Ile-X-X-Pro, obtained by comparing the affinity-isolated tryptic peptide with the affinity-isolated chymotryptic peptide and an overlapping tryptic peptide, shows homology with a sequence obtained from the NH2-terminal of a CNBr peptide from proteo glycan core protein of bovine nasal cartilage: Ser-Ser-Ala-Gly-Trp-Leu-Ala-Asp-Arg-Ser-Val-Arg-Tyr-Pro-Ile-Ser-. We suggest that the common sequence is structurally important to the function of these proteins and may be involved in the binding of both link protein and proteoglycan to hyaluronic acid.
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PMID:An amino acid sequence common to both cartilage proteoglycan and link protein. 390 75