Development of expression systems for the production of recombinant human serum albumin using the MOX promoter in Hansenula polymorpha DL-1

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Title
Development of expression systems for the production of recombinant human serum albumin using the MOX promoter in Hansenula polymorpha DL-1
Author(s)
Kang Hyun Ah; Whan Koo Kang; Won Kyuong Hong; Moo Woong Kim; Jeong Yoon Kim; Jung Hoon Sohn; Eui Sung Choi; Keun Bum Choe; Sang Ki Rhee
Bibliographic Citation
Biotechnology and Bioengineering, vol. 76, no. 2, pp. 175-185
Publication Year
2001
Abstract
To optimize the secretory expression of recombinant human serum albumin (HSA) under the control of methanol oxidase (MOX) promoter in the methylotrophic yeast Hansenula polymorpha DL-1, we analyzed several parameters affecting the expression of HSA from the MOX promoter. Removal of the 5′-untranslated region derived from HSA cDNA in the expression cassette led to at least a fivefold improvement of HSA expression efficiency at the translational level. With the optimized expression cassette, the gene dosage effect on HSA expression was abolished and thus, a single copy of the expression vector integrated into the MOX locus became sufficient for the maximal expression of HSA. Northern blot analysis revealed that the levels of HSA transcript did not increase any further upon increasing copy number. The mox-disrupted (moxΔ) transformant was constructed, in which the genomic MOX gene was transplaced with the HSA expression cassette, to examine the effect of the methanol oxidase-deficient phenotype of the host on HSA expression. The moxΔ transformant showed higher levels of HSA production in shake-flask cultures than the MOX wild-type transformant, especially at low concentrations of methanol and a twofold higher specific HSA production rate in fed-batch fermentation with an abrupt induction mode. The native prepro signal sequence of HSA secreted in H. polymorpha was correctly processed and the mature recombinant protein had a pl value identical to that of the authentic HSA. Our results suggest that the H. polymorpha expression systems developed in this study are suitable for large-scale production of recombinant albumin.
Keyword
human serum albuminhansenula polymorphaMOX promotercopy number5'-untranslated regionmethanol oxidase-deficient phenotype
ISSN
0006-3592
Publisher
Wiley
DOI
http://dx.doi.org/10.1002/bit.1157
Type
Article
Appears in Collections:
Division of Biomaterials Research > Synthetic Biology and Bioengineering Research Center > 1. Journal Articles
Division of Bio Technology Innovation > BioProcess Engineering Center > 1. Journal Articles
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