Characterization of glycosyl hydrolase family 3 beta-N-acetylglucosaminidases from Thermotoga maritima and Thermotoga neapolitana

Journal of Bioscience and Bioengineering
Kyoung-Hwa ChoiJaeho Cha

Abstract

The genes encoding beta-N-acetylglucosaminidase (nagA and cbsA) from Thermotoga maritima and Thermotoga neapolitana were cloned and expressed in Escherichia coli in order to investigate whether Thermotoga sp. is capable of utilizing chitin as a carbon source. NagA and CbsA were purified to homogeneity by HiTrap Q HP and Sephacryl S-200 HR column chromatography. Both enzymes were homodimers containing a family 3 glycoside hydrolase (GH3) catalytic domain, with a monomer molecular mass of 54 kDa. The optimal temperatures and pHs for the activities of the beta-N-acetylglucosaminidases were found to be 65-75 degrees C and 7.0-8.0, respectively. Both enzymes hydrolyzed chitooligomers such as di-N-acetylchitobiose and tri-N-acetylchitotriose, and synthetic substrates such as p-nitrophenyl-beta-D-glucose (pNPGlc), p-nitrophenyl N-acetyl beta-D-glucosamine (pNPGlcNAc), p-nitrophenyl di-N-acetyl beta-D-chitobiose (pNPGlcNAc(2)) and p-nitrophenyl tri-N-acetyl beta-D-chitotriose (pNPGlcNAc(3)). However, the enzymes had no activity against p-nitrophenyl-beta-D-galactose (pNPGal) and p-nitrophenyl N-acetyl beta-D-galactosamine (pNPGalNAc) or highly polymerized chitin. The k(cat) and K(m) values were determined for pNPGlcNAc, pNPGlcNAc(2) an...Continue Reading

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Citations

Jan 11, 2012·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Bo-Young YoonNam-Chul Ha
Jun 9, 2015·International Journal of Molecular Sciences·Nirakar PradhanAngelo Fontana
Jun 14, 2016·Extremophiles : Life Under Extreme Conditions·Junpei ZhouZunxi Huang
Nov 17, 2017·Applied Microbiology and Biotechnology·Rui ZhangZunxi Huang
Feb 7, 2013·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Hyung Ho Lee, Sang Taek Jung

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