Identification of a cDNA encoding an active asparaginyl endopeptidase of Schistosoma mansoni and its expression in Pichia pastoris 1

FEBS Letters

Caffrey, Conor R.²  Franklin, Christopher²  Salter, Jason P.²  McKerrow, James H.²  Sajid, Mohammed²  Mathieu, Mary A.²  Bogyo, Matthew¹  Lucas, Kimberley D.²  Gaffney, Alan M.² 

[1] Department of Biochemistry and Biophysics, UCSF, San Francisco, CA 94121, USA;Tropical Disease Research Unit at the Department of Pathology, UCSF, VAMC, 4150 Clement St.-113B, San Francisco, CA 94121, USA

关键词: Asparaginyl endopeptidase;    Legumain;    Sm32;    Recombinant expression;    Schistosoma;   

DOI  :  10.1016/S0014-5793(99)01798-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Asparaginyl endopeptidases, or legumains, are a recently identified family of cysteine-class endopeptidases. A single gene encoding a Schistosoma mansoni asparaginyl endopeptidase (a.k.a. Sm32 or schistosome legumain) has been reported, but by sequence homology it would be expected to yield an inactive product as the active site C197 had been replaced by N. We now describe a new S. mansoni gene in which C197 is present. Both gene products were expressed in Pichia pastoris. Autocatalytic processing to fully active C197 Sm32 occurred at acid pH. In contrast, N197 Sm32 was not processed and this is consistent with the hypothesis that C197 is essential for catalysis. This was confirmed by mutation of N197 to C and re-expression in Pichia. The availability of recombinant active Sm32 allows detailed analysis of its catalytic mechanism and its function(s) in the biology of this important human parasite.

【 授权许可】

Unknown