Zymogen of tadpole collagenase

Harper, Elvin; Bloch, Kurt J.; Gross, Jerome

doi:10.1021/bi00792a008

Cited by 172 publications

(59 citation statements)

References 28 publications

(12 reference statements)

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“…chains and the distribution of these chains within each type of collagen molecule. Electron microscopic studies of segment-long-spacing crystallites confirmed that the principal cleavage site of the cartilage collagen is located between bands 41 and 42 which is in agreement with that found for type I collagens using tadpole collagenase [25] or synovial collagenase [22]. We have also found that chick tendon collagen is cleaved by the synovial enzymc at thc same locus (unpublished results).…”

Section: Discussionsupporting

confidence: 84%

Action of Rheumatoid Synovial Collagenase on Cartilage Caollagen

Woolley

Glanville

Evanson

et al. 1975

European Journal of Biochemistry

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The action of purified rheumatoid synovial collagenase on purified cartilage collagen, U~( I I )~, in solution at 25 "C has been characterised. The enzyme attacked cartilage collagen in solution producing a 5 8 % reduction in specific viscosity and resulting in the appearance of two reaction products which represented approximately three-quarter and one-quarter fragments of the intact molecule as shown by disc electrophoresis in polyacrylamide gels containing sodium dodecyl sulphate. The a-chain fragments which comprised each of these components corresponded to molecular weights of approximately 74000 and 21 000.Electron microscopy of segment-long-spacing crystallites of the reaction products revealed three-quarter (TCA) and one-quarter (TCB) length fragments, and permitted accurate localization of the cleavage locus between bands41 and 42 (1-41). This cleavage site and the formation of TCA and TCB reaction products are very similar to those found for type-I collagen substrates.Cartilage collagen in solution was found to be more resistant to collagenase attack than tendon collagen, the rate of cartilage collagen degradation being six times slower than that for tendon collagen, as judged by viscometry. The mid-point melting temperatures (T,) for lathyritic cartilage and tendon collagen were 40.5 and 41.5 "C, and for the collagenase-produced reaction products 38.5 and 37.5 "C, respectively. The significance of these findings is discussed in relation to the structure of type I and I1 collagens.At 25 "C rheumatoid synovial collagenase, like other mammalian collagenases, has been shown to attack the collagen molecule in its native triple-helical conformation at a specific locus [l-31 yielding reaction products, commonly known as TCA and TCB, which represent three-quarter and one-quarter length fragments of the collagen molecule respectively. These products have been shown to be more susceptible than the native molecules to thermal denaturation [l]. Such findings were obtained using solutions of mammalian, type I, skin collagen having the molecular composition al(I)2a2, whereas collagen extracted from cartilage containing three identical a chains, and having a molecular composition al(II), [4-61, has been reported to be resistant to collagenases derived from gingival and polymorphonuclear leucocytes [7].Since earlier studies had shown that crude preparations of synovial collagenase were able to digest whole slices of articular cartilage [8,9] and more Enzyme. Collagenase (EC 3.4.24.3); trypsin (EC 3.4.21.4).Eur. J. Biochem. 50 (1975) recently, cartilage collagen has been shown to be attacked by rabbit corneal collagenase [lo] and human skin collagenase [l 13, we have compared the action of purified synovial collagenase on purified cartilage (type 11) and tendon (type I) collagens from lathyritic chicks. MATERIALS AND METHODS Preparation and Purification of Rheumatoid Synovial CollagenaseRheumatoid synovial tissue obtained at synovectomies was cut into fragments approximately 4 mm in greatest dimension and explante...

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Section: Discussionsupporting

confidence: 84%

Action of Rheumatoid Synovial Collagenase on Cartilage Caollagen

Woolley

Glanville

Evanson

et al. 1975

European Journal of Biochemistry

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“…Collagenase activity was assayed after trypsin activation of the medium (8) by the method of Harper et a1 (10). The details of this assay have been described recently (1 I).…”

Section: Methodsmentioning

confidence: 99%

Induction of the synthesis of latent collagenase and latent neutral protease in chondrocytes by a factor synthesized by activated macrophages

Ridge¹,

Oronsky²,

Kerwar³

1980

Arthritis & Rheumatism

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The spent medium of stimulated peritoneal macrophages coqtains a factor or factors that induce the de novo synthesis of collagenase and neutral protease(s) in primary cultures of articular chondrocytes. The synthesis of this inducing factor by stimulated macrophages is inhibited by cycloheximide, indicating that the factor is a protein. The inducing factor is heat stable and stable to tryspin treatment, but it is partially inactivated on treatment to pH 2.0. The inducing factor is not retained 04 concanavalin Sepharose, is staple to reductive alkylation, and has an apparent molecular weight of a p proximately 30,OOO. Addition of the inducing factor to cartilage slices also results in the synthesis of both collagenase and neutral protease by the slices. Both of these enzymes synthesized by chondrocytes are in the latent form and are activated by either trypsin or plasmin. The neutral protease (MW 22,000) is a metalloprotease and is inhibited by o-phenanthroline and by D-penicillamine but not by phenylmethane sulfonylfluoride or iodoacetic acid. Induction of these latent en- Submitted for publication August 6, 1979; accepted in revised form December 26, 1979. zymes in chondrocytes occurs between 8 and 16 hours after exposure to the inducing factor and is completely blocked by cycloheximide. The spent medium of stimulated lymphocytes (concanavalin A) does not induce the synthesis of these enzymes in chondrocytes, suggesting that the synthesis of the inducing factor may be a prop erty specific to stimulated macrophages.Immunologic disturbances leading to the infiltration of mononuclear and polymorphonuclear cells into the synovium are a characteristic feature of inflammatory joint disease (1). It has been postulated that these cells contribute to the breakdown of articular collagen and proteoglycans that is observed in the disease. This destruction of the matrix can occur by at least two mechanisms. In one mechanism, intracellular neutral proteases and collagenase of the polymorphonuclear cells are released in the synovium and lead to the enzymatic degradation of the cartilage matrix ( 2 4 ) . A second mechanism involves the interaction between the various cell types in the synovium leading to the synthesis of collagenase. Dayer et a1 (5,6) have shown that products synthesized by stimulated lymphocytes can activate synovial cells to synthesize abnormal levels of collagenase. Products of stimulated lymphocytes can also interact with macrophages and result in the synthesis of collagenase by the latter cells (7).Cell-cell interactions between mononuclear cells and chondrocytes have not been investigated in detail. In a recent preliminary communication by Phadke et a1

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“…Tadpole collagenase (EC 3.4.24.3) was purified as previously described (13), lyophilized, and stored at -20°until use. About 100 gg of collagenase were used to digest the procollagen and collagen extracted from three cranial bones; the digestion was carried out in 0.15 M NaCl, 50 mM Tris.…”

Section: Methodsmentioning

confidence: 99%

Interchain disulfide bonds in procollagen are located in a large nontriple-helical COOH-terminal domain.

Byers¹,

Click²,

Harper³

et al. 1975

Proc. Natl. Acad. Sci. U.S.A.

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Tadpole collagenase (EC 3.4.24.3) cleaved chick cranial bone procollagen into two triple-stranded fragments, PCA and PCB. Only pCB, with an estimated molecular weight of about 60,000 for each component chain after reduction, was found to contain interchain disulfide bonds. The analogous cleavage of collagen is known to produce a large NH2-terminal fragment with a molecular weight of 70 Since the initial identification of a biosynthetic precursor to collagen, it has become apparent that the nontriple-helical portion of the molecule is important for several functions, which may include molecular assembly, maintenance of intracellular solubility, transmembrane transport, and regulation of fibril formation (see ref. 1 for a review). Early structural studies of collagen precursors suggested, on the basis of electron micrographs of segment-long-spacing crystallites, that there was a single "noncollagenous" domain at the NH-terminus of each precursor chain (2, 3). Subsequent chemical studies of these precursors and of those procollagen-derived proteins which accumulated in the tissues of animals affected with dermatosparaxis tended to confirm the structural studies (4, 5).More recent studies of (a) peptides derived from procollagen secreted into culture medium by cells, (b) precursor chains protected from proteolysis during purification, and (c) peptides secreted into culture medium by cranial bone have suggested that the nonhelical, noncollagenous domain is considerably larger than was previously thought (6-9). Tanzer and his colleagues (10) have recently indicated that there is a noncollagenous peptide extension at the COOHterminus of the procollagen molecule, in addition to that at the NH2-terminus. In this report, we present data to show that the COOH-terminal domain of procollagen is larger than the NH2-terminal extensions, that the composition is different, and that interchain disulfide bonds occur exclusively in the COOH-terminal region. MATERIALS AND METHODSProcollagen, pulse labeled with L-[2,3-3H]proline (43.1 Ci/ mmol, New England Nuclear) for 18 min or with L-[G-3H]tryptophan (7.9 Ci/mmol, New England Nuclear) for 2 hr (7, 36) was extracted from 17-day-old chick embryo cranial bones with 1 M NaCl, 50 mM Trip-HCl, pH 7.5, to which 25 mM EDTA, 10 mM N-ethylmaleimide, 1 mM diisopropylfluorophosphate or phenylmethylsulfonylfluoride, and 1 mM benzamidine-HCl had been added to inhibit proteolysis. The bones were homogenized in that solvent, centrifuged at 39,000 X g for 15 min, and the labeled procollagen together with coextracted collagen was precipitated from the supernatant with 20% NaCl. The precipitate was collected by centrifugation, redissolved in extraction buffer, precipitated with 5% cold trichloroacetic acid and then dissolved in extraction buffer containing 0.1 M Tris-HCl, pH 7.5. Acid-extracted procollagen was obtained as previously described (4), except that extraction was limited to 1 hr, and was further purified by chromatography on DEAE-cellulose by minor modifications of the method of S...

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Zymogen of tadpole collagenase

Cited by 172 publications

References 28 publications

Action of Rheumatoid Synovial Collagenase on Cartilage Caollagen

Action of Rheumatoid Synovial Collagenase on Cartilage Caollagen

Induction of the synthesis of latent collagenase and latent neutral protease in chondrocytes by a factor synthesized by activated macrophages

Interchain disulfide bonds in procollagen are located in a large nontriple-helical COOH-terminal domain.

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