Expression of hepatitis B surface antigen S domain in recombinant Saccharomyces cerevisiae using GAL1 promoter.
Hepatitis B virus, a member of the hepadnavirus family, causes the acute and chronic diseases of the human liver. The S domain of hepatitis B virus surface antigen (sHBsAg) was expressed under the control of the galactose-inducible GAL1 promoter in recombinant Saccharomyces cerevisiae. Batch fermentation of S. cerevisiae 2805/pdeltaMFalpha-sHBsAg resulted in 4.92gl(-1) dry cell mass and 2.21mgl(-1) sHBsAg concentration. To improve the expression level of sHBsAg, the pdi1 gene encoding protein disulfide isomerase was coexpressed in recombinant S. cerevisiae. Batch fermentation of S. cerevisiae 2805/pdeltaMFalpha-sHBsAg+pPDI using 21gl(-1) glucose and 33gl(-1) galactose resulted in 9.75gl(-1) dry cell mass and 13.3mgl(-1) sHBsAg concentration, which were 2 and 6 times higher than those for S. cerevisiae 2805/pdeltaMFalpha-sHBsAg, respectively. Appearance of three sHBsAgs with different molecular sizes of 24kDa, 34kDa and 40kDa in immunoblot assay and their endoglycosidase treatment indicated that sHBsAg might be expressed in three types of the authentic, MFalpha signal sequence-containing and N-glycosylated MFalpha signal sequence-containing forms. Fed-batch fermentation of recombinant S. cerevisiae 2805 coexpressing the sHBsAg and pdi1 genes was carried out by feeding 600gl(-1) glucose continuously and controlling galactose concentration at around 20gl(-1). As a result, 20.2gl(-1) dry cell mass and 74.4mgl(-1) maximum sHBsAg concentration were obtained, which were 4.1 and 33.7 times higher than those for the batch fermentation of S. cerevisiae 2805/pdeltaMFalpha-sHBsAg, respectively.
Kim
Eun-Jeong
EJ
Department of Agricultural Biotechnology, Seoul National University, Seoul, Republic of Korea.
Park
Young-Kyoung
YK
Lim
Hyung-Kweon
HK
Park
Yong-Cheol
YC
Seo
Jin-Ho
JH
D000431:Ethanol / Q000378:metabolism; D005285:Fermentation; D005686:Galactokinase / Q000235:genetics*; D005690:Galactose / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006514:Hepatitis B Surface Antigens / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D019704:Protein Disulfide-Isomerases / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics*
D006514:Hepatitis B Surface Antigens; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D000431:Ethanol; C524219:GAL1 protein, S cerevisiae; D005686:Galactokinase; D019704:Protein Disulfide-Isomerases; D005947:Glucose; D005690:Galactose