Activity of select dehydrogenases with Sepharose-immobilized N-6-carboxymethyl-NAD

作者：Beauchamp Justin; Vieille Claire^*

来源：Bioengineered, 2015, 6(2): 106-110.

DOI：10.1080/21655979.2014.1004020

• analog
• catalysis
• cofactor
• dehydrogenase
• immobilize
• NAD
• sepharose

N-6-carboxymethyl-NAD (N-6-CM-NAD) can be used to immobilize NAD onto a substrate containing terminal primary amines. We previously immobilized N-6-CM-NAD onto sepharose beads and showed that Thermotoga maritima glycerol dehydrogenase could use the immobilized cofactor with cofactor recycling. We now show that Saccharomyces cerevisiae alcohol dehydrogenase, rabbit muscle L-lactate dehydrogenase (type XI), bovine liver L-glutamic dehydrogenase (type III), Leuconostoc mesenteroides glucose-6-phosphate dehydro-genase, and Thermotoga maritima mannitol dehydrogenase are active with soluble N-6-CM-NAD. The products of all enzymes but 6-phospho-D-glucono-1,5-lactone were formed when sepharose-immobilized N-6-CM-NAD was recycled by T. maritima glycerol dehydrogenase, indicating that N-6-immobilized NAD is suitable for use by a variety of different dehydrogenases. Observations of the enzyme active sites suggest that steric hindrance plays a greater role in limiting or allowing activity with the modified cofactor than do polarity and charge of the residues surrounding the N-6-amine group on NAD.

• 出版日期2015-4