Proteinase inhibitors of bovine nasal cartilage.
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ABSTRACT:
SUBMITTER: Roughley PJ
PROVIDER: S-EPMC1183848 | biostudies-other | 1978 Mar
REPOSITORIES: biostudies-other
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Project description:N-terminal analysis of aggrecan fragments lost from bovine nasal cartilage cultured in the presence of recombinant human interleukin 1alpha revealed a predominant ARGSVIL sequence with an additional ADLEX sequence. Production of the ARGSVIL-containing fragments has been attributed to the action of a putative proteinase, aggrecanase. The minor sequence (ADLEX) corresponds to a new reported cleavage product; comparison of this sequence with the available partial sequence of bovine aggrecan indicates that this is the product of a cleavage occurring towards the C-terminus of the protein. Matrix metalloproteinase (MMP) inhibitors inhibited aggrecan loss from bovine nasal explants incubated in the presence of recombinant human interleukin 1alpha. A strong correlation between inhibition of aggrecan metabolism and inhibition of stromelysin 1 (MMP 3) (r=0.93) suggests a role for stromelysin or a stromelysin-like enzyme in cartilage aggrecan metabolism. However, the compounds were approx. 1/1000 as potent in inhibiting aggrecan loss from the cartilage explants as they were in inhibiting stromelysin. There was little or no correlation between inhibition of aggrecan metabolism and inhibition of gelatinase B (MMP 9) or inhibition of collagenase 1 (MMP 1). Studies with collagenase inhibitors with a range of potencies showed a correlation between inhibition of collagenase activity and inhibition of collagen degradation in the cartilage explant assay. This indicates that in interleukin 1alpha-driven bovine nasal cartilage destruction, stromelysin (or a closely related enzyme) is involved in aggrecan metabolism, whereas collagenase is principally responsible for collagen degradation.
| S-EPMC1218345 | biostudies-other
Project description:1. Proteoglycans were extracted from bovine nasal cartilage with 2.0M-CaC2 or with 0.15M-KCl followed by 2.0M-CaC2.. Proteoglycan fractions were prepared from the extracts by density-gradient centrifugation in CsCl under 'associative' and 'dissociative' conditions. 2. The heterogeneity of the proteoglycan fractions was investigated by large-pore-gel electrophoresis. It was concluded that extracts made with 2.0M-CaCl2 or sequential 2.0M-CaCl2 contain two major species of proteoglycan 'subunit' of different hydrodynamic size, together with proteoglycan aggregates. Both 'subunits' have mobilities that are greater than those of proteoglycans obtained from pig articular cartilage McDevitt & Muir (1971) Anal. Biochem. 44, 612-622] and are therefore probably smaller in size than the latter. 3. Proteoglycan fractions isolated from cartilage extracted lith 0.15M-KCl separated into two main components on large-pore-gel electrophoresis with mobilities greater than those of proteoglycans extracted with 2.0M-CaCl2. Proteoglycans extracted at low ionic strength from bovine nasal cartilage are of similar hydrodynamic size to those extracted from pig articular cartilage under the same conditions [McDevitt & Muir (1971) Anal. Biochem. 44, 612-622]. 4. The role of endogenous proteolytic enzymes in producing proteoglycan heterogeneity, particularly in low-ionic-strength cartilage extracts is discussed. 5. Hyaluronic acid and 'link proteins' were present in the proteoglycan fraction separated from KCl extracts as well as in the fraction separated from CaCl2 extracts. Hyaluronic acid can only be identified in proteoglycan fractions by large-pore-gel electrophoresis after proteolysis and further purification of the fraction. 6. Collagen was extracted by both salt solutions and was tentatively identified as type II. Small amounts of collagen appear to be associated with the proteoglycan-aggregate fraction from the high-ionic-strength extract but not with the corresponding fraction from the KCl extract.
| S-EPMC1163861 | biostudies-other
Project description:1. The light fraction of the proteoglycan of bovine nasal cartilage was split by treatment with 0.1m-hydrochloric acid in acetone. The products were separated by gel filtration on 4% agarose and two retarded fractions were detected and isolated. These two fractions were found to have a Stokes radius of 134 and 47 A respectively, as determined by calibration of the column against proteins of known hydrodynamic volumes. 2. The 47 A fraction had a protein content of 4% and a glucosamine/galactosamine ratio 1:23. The 134 A fraction had a protein content of 20% and a glucosamine/galactosamine ratio 1:4.8. 3. The results of the viscometric studies on both fractions suggested that the 134 A fraction alone exhibited the property of undergoing reversible pH-dependent aggregation with a transition point at pH4.9. 4. It was concluded that these fractions could represent subunits of the native cartilage proteoglycan.
| S-EPMC1176692 | biostudies-other
Project description:1. Chondromucoprotein from bovine nasal cartilage was purified by cetylpyridinium chloride or by bismuth nitrate in acetone. 2. Amino acid compositions of crude and purified preparations were compared and few differences were found, in spite of the decrease in protein content on purification. 3. Amino acid analysis of bismuth-purified material revealed the existence of four groups of amino acids. Within each group, the amino acids were present in approximately equimolar concentrations. 4. Amino end-group assay on the same material showed six alpha-DNP derivatives. 5. A molecular weight of 6.3x10(5) for the protein-polysaccharide complex was calculated from the latter analysis.
| S-EPMC1198346 | biostudies-other
Project description:1. Three chondroitin sulphate components were isolated from adult bovine nasal cartilage after treatment with alkaline NaB3H. Average molecular weights of 13000, 18 600 and 28 000 were obtained for chondroitin sulphate species representing 10, 52 and 38% (w/w) of the total chondroitin sulphate respectively. Each chondroitin sulphate pool has a narrow molecular-weight distribution. 2. A proteoglycan subunit preparation, isolated from one nasal cartilage by extraction and density-gradient fractionation in dissociative solvents, partitioned on a CSCl density gradient according to size and composition. Variation of proteoglycan molecular weight across the gradient was directly related to the average chondrotin sulphate chain length, which in turn reflected the relative proportion of the three chondroitin sulphate pools in each proteoglycan fraction. Consideration of proteoglycan molecular parameters, compositions and behaviour on sedimentation leads to a proposal that nasal cartilage contains 3 distinct proteoglycan pools, each of which has a constant number of chondroitin sulphate side chains of different average molecular weight. 3. Molecular-weight distribution parameters for these proteoglycan preparations indicate that all serine residues on the protein core capable of initiating chondroitin sulphate biosynthesis are occupied and that proteoglycan polydispersity results directly from the polydispersity of the attached chondroitin sulphate component.
| S-EPMC1172406 | biostudies-other
Project description:Tryptic fragments of bovine nasal-cartilage proteoglycan, fractionated by dissociative density-gradient ultracentrifugation, were made to react by immunodiffusion against antiserum to a hyaluronidase-digest subfraction of cartilage proteoglycan monomer. This reaction produced two families of partly superimposed precipitin lines. One family was restricted to gradient fractions of medium or low buoyant density and included the immunoprecipitation reaction attributed to the hyaluronic acid-binding region of the cartilage proteoglycan monomer. The second family of precipitin lines was present alone in gradient fractions of high buoyant density. Immunodiffusion studies with antisera to relatively homogeneous keratan sulphate-rich and chondroitin sulphate-bearing fragment subfractions isolated from the gradient fraction of highest density indicated that both subfractions contained the antigenic determinants responsible for the second family of precipitin lines. Additional immunodiffusion studies, with the use of multispecific antisera to chondroitinase ABC digest and hyaluronidase digest of proteoglycan monomer, confirmed that the two subfractions shared antigenic determinants, and, in addition, indicated that these determinants were on one molecular species in the keratan sulphate-rich fragment subfraction and divided among at least three in the chondroitin sulphate-bearing fragment subfraction. Although an unprecedentedly large number of cartilage proteoglycan antigens could be recognized with the antisera employed in this cartilage proteoglycan antigens could be recognized with the antisera employed in this study, it was not possible to identify antigenic determinants unambiguously specific for the three structurally and functionally distinct regions of the cartilage proteoglycan monomer.
| S-EPMC1158285 | biostudies-other
Project description:Objective. Identify novel genes and pathways specific to superficial (SZ), middle (MZ) and deep zones (DZ) of normal articular cartilage. Methods. Articular cartilage was obtained from knees of 4 normal human donors. The cartilage zones were dissected on a microtome. RNA was analyzed on human genome arrays. Data obtained with human tissue were compared to bovine cartilage zone specific DNA arrays. Genes differentially expressed between zones were evaluated using direct annotation for structural or functional features, and by enrichment analysis for integrated pathways or functions. Results. The greatest differences were observed between SZ and DZ in both human and bovine cartilage. The MZ was transitional between the SZ and DZ and thereby shared some of the same pathways as well as structural/functional features of the adjacent zones. Cellular functions and biological processes enriched in the SZ relative to the DZ, include most prominently ECM receptor interactions, cell adhesion molecules, regulation of actin cytoskeleton, ribosome-related functions and signaling aspects such as Interferon gamma, IL4, CDC42Rac and Jak-Stat. Two pathways were enriched in the DZ relative to the SZ, including PPARG and EGFR_SMRTE. Conclusion. These differences in cartilage zonal gene expression identify new markers and pathways that govern the unique differentiation status of chondrocyte subpopulations.
2012-08-01 | GSE39796 | GEO
Project description:Objective. Identify novel genes and pathways specific to superficial (SZ), middle (MZ) and deep zones (DZ) of normal articular cartilage. Methods. Articular cartilage was obtained from knees of 4 normal human donors. The cartilage zones were dissected on a microtome. RNA was analyzed on human genome arrays. Data obtained with human tissue were compared to bovine cartilage zone specific DNA arrays. Genes differentially expressed between zones were evaluated using direct annotation for structural or functional features, and by enrichment analysis for integrated pathways or functions. Results. The greatest differences were observed between SZ and DZ in both human and bovine cartilage. The MZ was transitional between the SZ and DZ and thereby shared some of the same pathways as well as structural/functional features of the adjacent zones. Cellular functions and biological processes enriched in the SZ relative to the DZ, include most prominently ECM receptor interactions, cell adhesion molecules, regulation of actin cytoskeleton, ribosome-related functions and signaling aspects such as Interferon gamma, IL4, CDC42Rac and Jak-Stat. Two pathways were enriched in the DZ relative to the SZ, including PPARG and EGFR_SMRTE. Conclusion. These differences in cartilage zonal gene expression identify new markers and pathways that govern the unique differentiation status of chondrocyte subpopulations. 6 samples, 2 bovine donors, 3 conditions each donor (SZ, MZ and DZ), 0 donor replicates, comparisons made between SZ, MZ and DZ to identify differentially expressed genes.
2012-07-31 | E-GEOD-39796 | biostudies-arrayexpress
Project description:As there are more than 50 adenovirus serotypes, the likelihood of developing an effective vaccine is low. Here we describe inhibitors of the adenovirus proteinase (AVP) with the ultimate objective of developing anti-adenovirus agents. Inhibitors were identified via structure-based drug design using as druggable sites the active site and a conserved cofactor pocket in the crystal structures of AVP. A lead compound was identified that had an IC50 of 18 ?M. One of eight structural derivatives of the lead compound had an IC50 of 140 nM against AVP and an IC50 of 490 nM against the AVP with its cofactor bound.
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Project description:The Severe Acute Respiratory Syndrome (SARS) is a serious respiratory illness that has recently been reported in parts of Asia and Canada. In this study, we use molecular dynamics (MD) simulations and docking techniques to screen 29 approved and experimental drugs against the theoretical model of the SARS CoV proteinase as well as the experimental structure of the transmissible gastroenteritis virus (TGEV) proteinase. Our predictions indicate that existing HIV-1 protease inhibitors, L-700,417 for instance, have high binding affinities and may provide good starting points for designing SARS CoV proteinase inhibitors.
| S-EPMC7119134 | biostudies-literature