Preliminary Study on the Regulation of Mxr1 Phosphorylation by Ptp

doi:10.13560/j.cnki.biotech.bull.1985.2019-0020

Abstract

Abstract: The small molecule tyrosine phosphatase Ptp interacting with Pichia pastoris transcriptional activator Mxr1 was found by PULL-DOWN,and its function of dephosphorylation was verified,resulting in the hydrolysis of phosphate group in phosphorylated Mxr1 215th serine. Phosphorylation of Mxr1S215 in different media was detected by Western blot of Mxr1S215th specific site phosphorylation antibody. It was found that Mxr1in the wild strain was not phosphorylated in the glycerol medium,while phosphorylated in the methanol medium. In the Ptp-highly expressed strain,no phosphorylation occurred in Mxr1 in either glycerol or methanol medium,but phosphorylation was significantly higher in the Ptp-knockout strain than in the wild type one,indicating that Ptp indeed regulated Mxr1 phosphorylation. The expression in Escherichia coli was conducted,then the proteins were purified,and the Ptp enzymatic properties were determined. The P. pastorisMxr1S215A mutant strain was constructed,and several genes related to methanol metabolism were found to change at the transcriptional level,suggesting that it was regulated by Mxr1 phosphorylation at 215th site.

Key words: Pichia pastoris, transcriptionalactivator Mxr1, phosphatase, phosphorylation

HOU Cheng-lin, YANG Yan-kun, CHEN Jia-li, BAI Zhong-hu. Preliminary Study on the Regulation of Mxr1 Phosphorylation by Ptp[J]. Biotechnology Bulletin, 2019, 35(7): 108-113.

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