| Budget Amount *help |
¥6,300,000 (Direct Cost: ¥6,300,000)
Fiscal Year 1988: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 1987: ¥4,500,000 (Direct Cost: ¥4,500,000)
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| Research Abstract |
Achromobacter protease I (API) was classified into the mammalian-type serine proteoses from the results of the primary structural analysis of API. When the primary struature of API was compared with those of the other mammalian-type serine proteases, a few structural characteristics were found as follows ; (1) there are extensions of peptide chains in both N- and C-terminal regions of API ; (2) only Cys36-Cys58 is consevrative ; (3) Asp189 and Asp193, which are responsible for eatalytic activity, are replaced by Glu and Ser residues, respectively. In order to clarify the relationship between structure and fanction of API, the property and size of subsites of API were first investigated by using fluorogenic substrates. The result showed that API had three subsites,S1,S2,and S3 which corresponded to His213-Gly214-Gly214. This subsite sequence was different from the consensus subsite sequence, Ser-Trp(Gly)-Gly. So, it is suggested that the unique subsite sequence of API may be responsible for its paticular catalytic property.
On the other hand; we also tried identify a his residue of the catalytic site of API by using a new fluorogenic affinity-labeling reagent, DLCK. The result suggested that either His56 or His57 might be an active site his. Moreover, we investigated the contribution of amino groups of API to the structural stabilization of API by acetylation of the amino gruups. As the result, it was clarified that E-amino groups of Lys residues played an significant role in the structural stabilization of a
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