Interaction of cysteine proteinases with recombinant kininogen domain 2, expressed in Escherichia coli.
The calpain-binding domain 2 of the kininogens, the major plasma inhibitors of cysteine proteinases, was expressed in Escherichia coli. Expression of soluble protein was optimal at 15 degrees C and was augmented by growing the bacteria in sorbitol and betaine. The recombinant domain showed high affinity (Ki 0.3-1 nM) for cathepsin L and papain, and a somewhat lower affinity (Ki approximately 15 nM) for calpain. The binding to cathepsin H was substantially weaker, and no inhibition of actinidin and cathepsin B was detected. The affinity for cathepsin L was comparable to that reported for the domain isolated from plasma L-kininogen, whereas the affinities for papain and calpain were about tenfold lower. The latter difference may be due to the recombinant domain being nonglycosylated.
Department of Veterinary Medical Chemistry, Swedish University of Agricultural Sciences, Uppsala Biomedical Center.
K
K
Ylinenjärvi
T W
TW
Prasthofer
N C
NC
Martin
I
I
Björk
D000818:Animals; D056668:Cathepsin L; D002403:Cathepsins / Q000378:metabolism*; D002417:Cattle; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D010450:Endopeptidases; D004926:Escherichia coli; D006801:Humans; D007704:Kininogens / Q000235:genetics / Q000378:metabolism*; D010206:Papain / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012756:Sheep
D007704:Kininogens; D011994:Recombinant Proteins; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C534281:CTSL protein, human; D056668:Cathepsin L; C534280:Ctsl protein, rat; D010206:Papain