期刊
PROTEIN SCIENCE
卷 9, 期 12, 页码 2567-2572出版社
CAMBRIDGE UNIV PRESS
DOI: 10.1110/ps.9.12.2567
关键词
cathepsin L; Fasciola hepatica; homology modeling; S-2 subsite specificity
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.
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