Collagenolytic serine proteinase from Euphausia superba Dana (Antarctic krill).
1. A serine proteinase isolated from E. superba shows collagenolytic properties: it acts on collagens from Achilles tendon (type I and V) and reconstituted fibrils of calf skin collagen under conditions that do not denature the substrates. 2. At 25 degrees C and pH 7.5 the enzyme both splits the calf skin collagen in solution to the fragments TCA and TCB and catalyses the conversion of dimeric molecules to monomeric chains. 3. The enzyme exhibits strong chymotrypsin-like and lower trypsin-like activities. 4. All the enzyme activities are inhibited to the same degree by diisopropylfluorophosphate (DFP), phenylmethylsulphonyl fluoride (PMSF), N alpha-tosyl-L-lysine chloromethyl ketone (TLCK), soybean trypsin inhibitor (SBTI), chicken ovomucoid (CHOM), chymostatin and leupeptin. None of the activities is inhibited by chelating agents and L-cysteine. 5. pH-Optima of the proteinase in protein substrates hydrolysis (6.0-6.2) are lower than those of synthetic substrates cleavage (7.8-8.0 in the case of BzTyrOEt and 8.7-8.9 for BzArgOEt). 6. Four from nine cysteine residues present in the enzyme molecule possess free thiol-groups. Since the enzyme is inhibited by p-chloromercuribenzoate (pCMB), N-ethylmaleimide (NEM) and iodoacetic acid (IAA), the role of its thiol-groups has been discussed.
Turkiewicz
M
M
Institute of Technical Biochemistry, Technical University of Lódź, Poland.
Galas
E
E
Kalinowska
H
H
D000818:Animals; D000864:Antarctic Regions; D002417:Cattle; D002845:Chromatography; D002848:Chromatography, DEAE-Cellulose; D002852:Chromatography, Ion Exchange; D003094:Collagen / Q000378:metabolism; D003445:Crustacea / Q000201:enzymology*; D017886:Durapatite; D006801:Humans; D006882:Hydroxyapatites; D007700:Kinetics; D003012:Microbial Collagenase / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D012867:Skin; D013379:Substrate Specificity
D006882:Hydroxyapatites; D003094:Collagen; D017886:Durapatite; D012697:Serine Endopeptidases; D003012:Microbial Collagenase