Modification of Strong Promoter PCAT1 of Pichia pastoris and Analysis of Transcription Factor Binding Sites
Article
Figures
Metrics
Preview PDF
Reference
Related
Cited by
Materials
Abstract:
In this study, the methanol-inducible strong promoter PCAT1 of Pichia pastoris was modified by random mutation, characterized by green fluorescent protein. After high-throughput screening, a PCAT1 promoter library with an activity range of 25.57%- 138.12% was obtained. It was found that the PCAT1 variant 6-39 whose activity increased by 38.12% was mutated from A to G at the -118 bp in the 5 '-3'direction.The online prediction software for transcription factor binding sites, MatInspector, was used to analyze the change of transcription factor binding sites of the PCAT1 variants. Compared to wild-type PCAT1, the binding site of F$YMCM of the PCAT1 variant was reduced, and the binding sites of F$CSRE and F$YGAL increased. Then, the transcription factor binding sites of 16 promoters of the genes involved in methanol utilization pathway of Pichia pastoris were analyzed by MatInspector. It was found that the binding sites of the transcription factor, F$CSRE, were widely distributed in the methanol-inducible promoters, whilst the binding sites of F$YGAL were extensively distributed only in the constitutive promoter. Thus, the increase of the binding sites of F$CSRE is most likely to increase the methanol-inducing activity of PCAT1, which provides a reference for the analysis of the key sites of the promoters of the genes for the methanol utilization pathway of Pichia pastoris.
