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For the record: A single amino acid substitution affects substrate specificity in cysteine proteinases from Fasciola hepatica
Author(s) -
Smooker Peter M.,
Whisstock James C.,
Irving James A.,
Siyaguna Sugeeshwara,
Spithill Terry W.,
Pike Robert N.
Publication year - 2000
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.9.12.2567
Subject(s) - cathepsin o , fasciola hepatica , biochemistry , cathepsin l1 , cathepsin e , cathepsin , cathepsin c , cathepsin l , proteases , biology , cathepsin h , cathepsin s , cathepsin a , cathepsin b , amino acid , cysteine protease , peptide sequence , cysteine , microbiology and biotechnology , recombinant dna , enzyme , gene , zoology , helminths
The trematode Fasciola hepatica secretes a number of cathepsin L‐like proteases that are proposed to be involved in feeding, migration, and immune evasion by the parasite. To date, six full cDNA sequences encoding cathepsin L preproproteins have been identified. Previous studies have demonstrated that one of these cathepsins (L2) is unusual in that it is able to cleave substrates with a proline in the P 2 position, translating into an unusual ability (for a cysteine proteinase) to clot fibrinogen. In this study, we report the sequence of a novel cathepsin (L5) and compare the substrate specificity of a recombinant enzyme with that of recombinant cathepsin L2. Despite sharing 80% sequence identity with cathepsin L2, cathepsin L5 does not exhibit substantial catalytic activity against substrates containing proline in the P 2 position. Molecular modeling studies suggested that a single amino acid change (L69Y) in the mature proteinases may account for the difference in specificity at the S 2 subsite. Recombinant cathepsin L5/L69Y was expressed in yeast and a substantial increase in the ability of this variant to accommodate substrates with a proline residue in the P 2 position was observed. Thus, we have identified a single amino acid substitution that can substantially influence the architecture of the S 2 subsite of F. hepatica cathepsin L proteases.