Expression of an alpha-galactosidase gene under control of the homologous inulinase promoter in Kluyveromyces marxianus.
For expression of the alpha-galactosidase gene from Cyamopsis tetragonoloba in Kluyveromyces marxianus CBS 6556 we have used the promoter of the homologous inulinase-encoding gene (INU1). The INU1 gene has been cloned and sequenced and the coding region shows an identity of 59% with the Saccharomyces cerevisiae invertase gene (SUC2). In the 5'-flanking region of INU1 we found a sequence (TAAATCCGGGG) that perfectly matches to the MIG1 binding consensus sequence (WWWWTSYGGGG) of the S. cerevisiae GAL1, GAL4 and SUC2 genes. Using the K. marxianus INU1 promoter and prepro-signal sequence, we obtained a high alpha-galactosidase production level (153 mg/l) and a secretion efficiency of 99%. Both the production level and the secretion efficiency were significantly reduced when the INU1 pro-peptide was deleted. With either the S. cerevisiae PGK or GAL7 promoter we could obtain only low alpha-galactosidase production levels (2 mg/l).
Bergkamp
R J
RJ
Laboratorium voor Biochemie en Moleculaire Biologie, Vrije Universiteit, Amsterdam, The Netherlands.
Bootsman
T C
TC
Toschka
H Y
HY
Mooren
A T
AT
Kox
L
L
Verbakel
J M
JM
Geerse
R H
RH
Planta
R J
RJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D015870:Gene Expression / Q000502:physiology; D005800:Genes, Fungal / Q000235:genetics / Q000502:physiology*; D006026:Glycoside Hydrolases / Q000235:genetics*; D007716:Kluyveromyces / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D000519:alpha-Galactosidase / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
D006026:Glycoside Hydrolases; D000519:alpha-Galactosidase; C023086:inulinase