Satoshi Mitsuda

Studies on enantioselective hydrolysis of the acetic ester of a secondary alcohol with Arthrobacter lipase

SummaryCharacteristics of the enantioselective hydrolysis of the acetic ester of 4-hydroxy-3-methyl-2-(2-propynyl)-2-cyclopentenone (HMPC) by Arthrobacter lipase were investigated in a water/oil biphasic reaction mixture. Kinetic studies revealed that the strict enantioselectivity was entirely due to a difference in the catalytic constants for the enantiomeric substrates and that (S)-HMPC acetate acted as a competitive inhibitor. The comparison of enantioselectivity for the acetates of HMPC analogues indicated that hydrophobic substituents in the HMPC molecule were essential for the strict enantioselectivity.

Factors affecting the production of nitrile hydratase by thermophilic Bacillus smithii SC-J05-1.

Ammonium present in the medium was found to control the production of nitrile hydratases (NHase) by a moderate thermophile, Bacillus smithii SC-J05-1. The enzyme production is initiated in the absence of ammonium and suppressed in the presence of ammonium. In addition to cobalt ions as the coordinated metal ion, manganese ions are also required for NHase production.

Characterization of a unique scrambled peptide derived from the CD4 CDR3-related region which shows substantial activity for blocking HIV-1 infection

We have previously identified CD4 peptides that exhibited blocking activity on human immunodeficiency virus type 1 (HIV-1) infection, i.e. CD4(68-130) and CD4(66-92) which include the region corresponding to the third complementarity-determining region of IgG. Here we describe a unique peptide derived from CD4(66-92), altered in amino acid sequence but not in composition, which was found to have increased anti-HIV-1 activity. The acidic amino acid residues in this scrambled peptide, S1, localized at the N-terminus, while in the native peptide they clustered at the C-terminus. On the other hand, a second scrambled peptide, S2, in which the acidic amino acid residues were fully dispersed, did not show any anti-HIV-1 activity. However, we could not identify any correlation between CD4(66-92) and S1 peptides by their hydrophobic or circular dichroism spectrum analyses. The results provide insight into the mechanisms of HIV-1 gp120 and CD4 interaction and may be useful as a new approach to AIDS therapy.

Cloning, purification, crystallization and preliminary X-ray diffraction analysis of nitrile hydratase from the themophilic Bacillus smithii SC-­J05-1

Nitrile hydratase (NHase) converts nitriles to the corresponding amides and is recognized as having important industrial applications. Purification, cloning, crystallization and initial crystallographic studies of the NHase from Bacillus smithii SC-J05-1 (Bs NHase) were conducted to analyze the activity, specificity and thermal stability of this hydrolytic enzyme. Bs NHase was purified to homogeneity from microbial cells of B. smithii SC-J05-1 and the nucleotide sequences of both the alpha- and beta-subunits were determined. Purified Bs NHase was used for crystallization and several crystal forms were obtained by the vapour-diffusion method. Microseeding and the addition of magnesium ions were essential components to obtain crystals suitable for X-ray diffraction analysis.