Title of article :
Enzymatic synthesis of nylon-6 units in organic solvents containing low concentrations of water
Author/Authors :
Kawashima، نويسنده , , Yasuyuki and Yasuhira، نويسنده , , Kengo and Shibata، نويسنده , , Naoki and Matsuura، نويسنده , , Yusuke and Tanaka، نويسنده , , Yusuke and Taniguchi، نويسنده , , Masaaki and Miyoshi، نويسنده , , Yoshiaki and Takeo، نويسنده , , Masahiro and Kato، نويسنده , , Dai-ichiro and Higuchi، نويسنده , , Yoshiki and Negoro، نويسنده , , Seiji، نويسنده ,
Issue Information :
روزنامه با شماره پیاپی سال 2010
Pages :
8
From page :
81
To page :
88
Abstract :
NylB′ carboxylesterase, which is 88% homologous to functional 6-aminohexanoate-dimer hydrolase (NylB) from Arthrobacter sp., possesses trace synthetic activity [0.0004 μmol min−1 mg−1 (U/mg)] from 6-aminohexanoate (Ahx) to its oligomers in 90% tert-butyl alcohol. The synthetic activity and the ratio of the synthetic activity to the hydrolytic activity were significantly affected by amino acid substitutions at positions 181, 266 and 370. The synthetic activity was enhanced to 2.7 U/mg by G181D-H266N substitutions, and the activity was further enhanced in the G181D-H266N-D370Y triple mutant to a level approximately 104-fold greater than the parental carboxylesterase form (3.4 U/mg), which was nearly equal to the ordinary hydrolytic activity in water (type A-mutants). Type A-mutants possessed more than 50% of the 6-aminohexanoate-linear dimer (Ald)-hydrolytic activity at 0–70% tert-butyl alcohol, but the synthetic reaction became predominant at 85–90% tert-butyl alcohol. In contrast, type B-mutants (G181E-H266N and G181N-H266N) possessed quite low levels of Ald-hydrolytic activity (<0.01 U/mg) at 0–70% tert-butyl alcohol. However, both the hydrolytic and synthetic activities were enhanced at higher concentrations, and the maximum activity was obtained at 90% tert-butyl alcohol for both hydrolysis and synthesis. In a type C-mutant (R187S-F264C-D370Y), the Ald-hydrolytic activity was enhanced to approximately 80-fold that of the parental carboxylesterase, but the mutant barely demonstrated any synthetic activity. On the basis of the three-dimensional structure of the Ald-bound enzyme and a kinetic study for typical mutant enzymes, we propose that the efficient enzymatic syntheses of nylon-6 units were achieved by (i) stable binding of the 1st-Ahx at the N-terminal region with Asp181, (ii) interaction of the 2nd-Ahx at the C-terminal region with Tyr370, and (iii) motion of Tyr170 that generated a closed form in the catalytic center of Ald hydrolase.
Keywords :
Unnatural amide compound , Enzymatic synthesis , ?-lactamase , 6-aminohexanoate-dimer hydrolase , carboxylesterase , nylon oligomer
Journal title :
Journal of Molecular Catalysis B Enzymatic
Serial Year :
2010
Journal title :
Journal of Molecular Catalysis B Enzymatic
Record number :
1714508
Link To Document :
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