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Selective expression of nonsecreted triple-helical and secreted single-chain recombinant collagen fragments in the yeast Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
25658 43588 2006 9 PDF Available
Title
Selective expression of nonsecreted triple-helical and secreted single-chain recombinant collagen fragments in the yeast Pichia pastoris
Abstract

High-level recombinant expression systems for the production of stable triple-helical human collagens and collagen fragments have been developed in the yeast Pichia pastoris. Collagen fragments are secreted as single-chain polypeptides by the yeast α-mating factor pre–pro sequence, but secretion of full-length triple-helical procollagen molecules has not been achieved despite the use of the same secretory signal. We studied here the effects of the secretory signal and the conformation and size of the collagen polypeptide on its secretion in P. pastoris. Unlike the collagen signal sequence, the α-mating factor pre–pro sequence led to efficient secretion of single-chain 45 and 9 kDa type I collagen fragments. The efficiency was dependent on the length of the collagen polypeptide, as secretion of single-chain full-length 90 kDa α1(I) polypeptides was less efficient than that of the 45 kDa fragment. Furthermore, the conformation of the collagen polypeptides had a marked effect on secretion, as induction of trimerization of the 45 and 9 kDa fragments by either the C propeptide or the small trimerizing domain foldon led to an accumulation of triple-helical molecules inside the cells despite the presence of the α-mating factor pre–pro sequence. Our results show that P. pastoris is a suitable host for the development of tailored expression systems aimed at selective production of nonsecreted triple-helical and secreted single-chain collagen fragments of varying lengths for specific purposes.

Keywords
Collagen; Gelatin; Yeast; Secretion
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Selective expression of nonsecreted triple-helical and secreted single-chain recombinant collagen fragments in the yeast Pichia pastoris
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 123, Issue 2, 17 May 2006, Pages 248–256
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us