Kinetic and Circular Dichroism Spectroscopic Comparison among Three Forms of Glucoamylase from a Rhizopus sp.

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Three forms of glucoamylase [EC 3. 2. 1. 3] of a Rhizopus sp., Gluc_1 (M. W. 74000), Gluc_2 (M. W. 58600) and Gluc_3 (M. W. 61400), were compared by circular dichroism (CD) spectroscopy and kinetic studies. The CD spectra of the enzymes indicated that Gluc_2 and Gluc_3. which lack fragments from the N-terminal part of Gluc_1,resembled Gluc_1 in protein backbone conformation, although with some differences in the states of aromatic amino acid side chains. Therefore, the N-terminal part of Gluc_1 appears not to have a critical effect on the gross conformation of the remaining domain of Gluc_1,though two glycopeptides, presumably released from the N-terminal part of Gluc_1,fragments H (M. W. 16700) and L (M. W. (14400), had different conformations from that of Gluc_1. The three enzymes each contained 1 SH in a buried form, as well as 1 S-S bridge, and had similar susceptibility to denaturants. On the other hand, Gluc_2 and Gluc_3 differed markedly from Gluc_1 in the K_m values for large substrates, but differed little in the K_m and V_<max> values for small substrates or in the V_<max> values for large substrates, except for pullulan, which is highly branched. Gluc_1,but not Gluc_2 or Gluc_3,may have an additional site (s) interacting with large substrates, besides the active center.
社団法人日本薬学会の論文
1985-01-25