Journal of Microbiology and Biotechnology
The Korean Society for Microbiology and Biotechnology publishes the Journal of Microbiology and Biotechnology.

2014 ; Vol.24-3: 337~345

AuthorNagaraj Govindappa, Manjunatha Hanumanthappa, Krishna Venkatarangaiah, Sankar Periyasamy, Suma Sreenivas, Rajeev Soni, Kedarnath Sastry
Place of dutyBiocon Research Limited - SEZ Unit, Bengaluru 560 099, India
TitleA New Signal Sequence for Recombinant Protein Secretion in Pichia pastoris
PublicationInfo J. Microbiol. Biotechnol.2014 ; Vol.24-3
AbstractPichia pastoris is one of the most widely used expression systems for the secretory expression of recombinant proteins. The secretory expression in P. pastoris usually makes use of the prepro MATα sequence from Saccharomyces cerevisiae, which has a dibasic amino acid cleavage site at the end of the signal sequence. This is efficiently processed by Kex2 protease, resulting in the secretion of high levels of proteins to the medium. However, the proteins that are having the internal accessible dibasic amino acids such as KR and RR in the coding region cannot be expressed using this signal sequence, as the protein will be fragmented. We have identified a new signal sequence of 18 amino acids from a P. pastoris protein that can secrete proteins to the medium efficiently. The PMT1-gene-inactivated P. pastoris strain secretes a ~30 kDa protein into the extracellular medium. We have identified this protein by determining its N-terminal amino acid sequence. The protein secreted has four DDDK concatameric internal repeats. This protein was not secreted in the wild-type P. pastoris under normal culture conditions. We show that the 18-amino-acid signal peptide at the N-terminal of this protein is useful for secretion of heterologous proteins in Pichia.
Full-Text
Key_wordDDDK protein, PMT1 gene inactivation, P. pastoris strain BICC 9450, Protein secretion
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