Purification and characterization of heterologously expressed nitrilases from filamentous fungi

0377497 - MBÚ 2013 RIV DE eng J - Článek v odborném periodiku
Petříčková, Alena - Veselá, Alicja Barbara - Kaplan, Ondřej - Kubáč, David - Uhnáková, Bronislava - Malandra, A. - Felsberg, Jürgen - Rinágelová, Anna - Weyrauch, P. - Křen, Vladimír - Bezouška, Karel - Martínková, Ludmila
Purification and characterization of heterologously expressed nitrilases from filamentous fungi.
Applied Microbiology and Biotechnology. Roč. 93, č. 4 (2012), s. 1553-1561. ISSN 0175-7598. E-ISSN 1432-0614
Grant CEP: GA ČR(CZ) GAP504/11/0394; GA ČR GD305/09/H008; GA AV ČR IAA500200708; GA MŠMT(CZ) LC06010; GA MŠMT OC09046
Výzkumný záměr: CEZ:AV0Z50200510
Klíčová slova: Chaperones * Nitrilase * Aspergillus niger
Kód oboru RIV: CE - Biochemie
Impakt faktor: 3.689, rok: 2012

Nitrilases from Aspergillus niger CBS 513.88, A. niger K10, Gibberella moniliformis, Neurospora crassa OR74A, and Penicillium marneffei ATCC 18224 were expressed in Escherichia coli BL21-Gold (DE3) after IPTG induction. N. crassa nitrilase exhibited the highest yield of 69,000 UL−1 culture. Co-expression of chaperones (GroEL/ES in G. moniliformis and P. marneffei; GroEL/ES and trigger factor in N. crassa and A. niger CBS 513.88) enhanced the enzyme solubility. Specific activities of strains expressing the former two enzymes increased approximately fourfold upon co-expression of GroEL/ES. The enzyme from G. moniliformis (co-purified with GroEL) preferred benzonitrile as substrate (Km of 0.41 mM, Vmax of 9.7 μmol min−1 mg−1 protein). The P.marneffei enzyme (unstable in its purified state) exhibited the highest Vmax of 7.3 μmol min−1 mg−1 protein in cellfree extract, but also a high Km of 15.4 mM, for 4-cyanopyridine
Trvalý link: http://hdl.handle.net/11104/0209640