Synthetic substrates for vertebrate collagenase

Masui, Yoshihiro; Takemoto, Takashi; Sakakibara, Shumpei; Hori, Hisae; Nagai, Yutaka

doi:10.1016/0006-2944(77)90026-6

Cited by 150 publications

(78 citation statements)

References 3 publications

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“…One unit of enzyme degraded 1 ,ug of gelatin/min at 37 'C. Peptidolytic activity was determined by proteolytic degradation of the synthetic octapeptide (dinitrophenyl-Pro-Gln-Gly-Ile-Ala-Gly-Gln-D-Arg) (DNP-peptide) as described by Masui et al (1977). One unit of enzyme degraded 1 ,uM substrate/min at 37°C.…”

Section: Materials and Methods Materialsmentioning

confidence: 99%

Fragmentation of human polymorphonuclear-leucocyte collagenase

Knauper

Osthues

DeClerck

et al. 1993

Biochemical Journal

110

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Human polymorphonuclear-leucocyte collagenase (M(r) 64,000) shows autoproteolytic degradation to two major fragments of M(r) 40,000 and M(r) 27,000. N-terminal sequence data and investigation of the substrate specificity of the fragments demonstrate that the M(r)-40,000 fragment corresponds to the catalytic domain, whereas the M(r0-27,000 fragment shows no enzymic activity. The activity profile of the M(r)-40,000 fragment is comparable with the specificity of the intact active collagenase (M(r) 64,000), but the ability to cleave collagen was lost. The enzymic activity of this fragment can be inhibited by either tissue inhibitor of metalloproteinase (TIMP)-1 or recombinant TIMP-2 in a 1:1 molar ratio. The C-terminal part of the enzyme (M(r) 27,000), important for the binding reaction with collagen substrates, is involved in collagenolysis.

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Section: Materials and Methods Materialsmentioning

confidence: 99%

Fragmentation of human polymorphonuclear-leucocyte collagenase

Knauper

Osthues

DeClerck

et al. 1993

Biochemical Journal

110

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“…Enzymic activity was routinely determined by degradation of the synthetic octapeptide dinitrophenyl (Dnp)-Pro-Gln-Gly-Ile-AlaGly-Gln-D-Arg-OH (Masui et al, 1977), whereas collagenolytic activity was determined by degradation of soluble type I collagen and analysis by SDS/PAGE followed by Coomassie Blue staining and gel scanning (Welgus et al, 1981). One unit of peptidolytic activity corresponded to the degradation of 1 ,umol of substrate per min at 37 'C.…”

Section: Determination Of Peptidolytic and Collagenolytic Activitymentioning

confidence: 99%

Direct activation of human neutrophil procollagenase by recombinant stromelysin

Knäuper

Wilhelm

Seperack

et al. 1993

Biochemical Journal

149

105

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Human neutrophil procollagenase was activated by incubation with recombinant active stromelysin. Activation was achieved by cleavage of the Gly78-Phe79 peptide bond at the end of the propeptide domain in a single-step activation mechanism. In addition, accelerated activation was achieved when N-terminally truncated, latent collagenase (with Phe49 as its N-terminal residue) was incubated with recombinant active stromelysin. Determination of the specific activity of recombinant-stromelysin-activated neutrophil collagenase with dinitrophenyl-octapeptide or type I collagen demonstrated the generation of high specific activity. The specific activity of stromelysin-activated enzyme was considerably higher than that of trypsin- or HgCl2-activated collagenase. Thus human neutrophil collagenase is superactivated, like the homologous fibroblast collagenase [Murphy, Cockett, Stephens, Smith and Docherty (1987) Biochem. J. 248, 265-268]. The occurrence of Phe79 at the N-terminus of the neutrophil collagenase seemed to be critical for superactivation, which is in agreement with data published by Suzuki, Enghild, Morodomi, Salvesen and Nagase [(1990) Biochemistry 29, 10261-10270] on fibroblast collagenase.

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“…General matrix metalloproteinase-activities were assayed with the synthetic substrate dinitrophenyl-Pro-Gln-Gly-Ile-Ala-Gly-Gln-DArg (dinitrophenyl-labelled peptide, 250 mg/1) as described by Masui et al (22) and with the fluorogenic substrate (7-methoxycoumarin-4-yl)acetyl-Pro-Leu-Gly-Leu[3-(2,4-dinitrophenyl)-2,3-diaminopropionyl]-Ala-Arg-NH 2 (25 umol/l) as introduced by Knight et al (23).…”

Section: Enzyme Assaysmentioning

confidence: 99%

“…The media were ultrafiltrated at a relative molecular mass cut-off of Μ τ 30000 to eliminate TIMP-1. The gelatinase A-TIMP-2-containing fractions were incubated with active gelatinase Β for 30 min at 37 °C and the activity of gelatinase B was measured using the dinitrophenyl-labelled peptide (22) as substrate (incubation: 30 min, 37 °C).…”

Section: Enzyme Assaysmentioning

confidence: 99%

Substance P Induces the Secretion of Gelatinase A from Human Synovial Fibroblasts

Hecker-Kia

Kolkenbrock²,

Orgel³

et al. 1997

Clinical Chemistry and Laboratory Medicine

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Summary: We investigated the secretion of the matrix metalloproteinases, interstitial collagenase (matrix metalloproteinase-1), gelatinase A (matrix metalloproteinase-2) and stromelysin-1 (matrix metalloproteinase-3) in human synovial fibroblasts after stimulation with the neuropeptide substance P.Human synovial fibroblasts were stimulated with substance P or interleukin-lß (IL-lß). In the cell culture media gelatinase A, interstitial collagenase and stromelysin-1 were identified and their activities towards different substrates were determined.Substance P in synovial fibroblasts induced an increase in the overall matrix metalloproteinase activity towards the dinitrophenyl-labelled peptide by 85%, against an increase of 124% after stimulation with IL-lß. In case of substance P stimulation, the increase in activity reflects a significantly enhanced secretion of gelatinase A, whereas no significant increase of stromelysin-1 and collagenase secretion could be observed.The matrix metalloproteinase pattern showing the highest gelatinase A secretion was obtained after stimulation with substance P. This pattern was very pronounced and differed very clearly from the pattern seen after IL-lß stimulation which caused a significant rise in collagenase and stromelysin-1 activity. We assume that distinct stimulation pathways are involved and that the neuropeptide (substance P), which is always present in the inflamed joint, plays its own and separate role in proliferative processes leading to the cartilage destruction.Introduction broblasts (5, 11). In these cells, but also in monocytes/

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Synthetic substrates for vertebrate collagenase

Cited by 150 publications

References 3 publications

Fragmentation of human polymorphonuclear-leucocyte collagenase

Fragmentation of human polymorphonuclear-leucocyte collagenase

Direct activation of human neutrophil procollagenase by recombinant stromelysin

Substance P Induces the Secretion of Gelatinase A from Human Synovial Fibroblasts

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