Characterization of UGPase from Aureobasidium pullulans NRRL Y-12974 and Application in Enhanced Pullulan Production.

Li, Haifeng; Zhang, Yunyi; Gao, Yunyun; Lan, Yuanyang; Yin, Xiaopu; Huang, Lifeng

UDPG pyrophosphatase (UGPase) plays an important role in carbohydrate metabolism, catalyzing a reversible production of uridine diphosphate glucose (UDPG) and pyrophosphate (PPi) from Glc-1-P and UTP. UGPase gene from Aureobasidium pullulans NRRL Y-12974 was cloned, overexpressed in Escherichia coli. The recombinant UGPase possess molecular mass of 55 KDa and specific activity of 7.33 U/mg protein. The K values of rUGPase were 5.045 μM against UTP and 3.333 μM against Glc-1-P. The V values of rUGPase were 3.467 μM minagainst UTP and 2.817 μM min against Glc-1-P. And, it does not catalyze Glc-1-P and ATP, nor galactose-1-P and UTP. Homolgous expression of UGPase in native organism can improve the intracellular UDPG concentration by 4.7-fold time. The yield of pullulan in engineering strain A4 was improved to 18.2 g g cell dry weight which is 1.3-fold time of parent strain. No obvious change of growth was found between engineering strain and parent strain. To the best of our knowledge, this is the first report of improving pullulan yield in A. pullulans using metabolic engineering technique.

AUREOBASIDIUM pullulans; INORGANIC pyrophosphatase; EXPRESSIVE behavior; PULLULANASE; URIDINE diphosphate sugars

Applied Biochemistry & Biotechnology, 2016, Vol 178, Issue 6, p1141

0273-2289

Academic Journal

10.1007/s12010-015-1934-2