Sequencing and functional analysis of the Hansenula polymorpha genomic fragment containing the YPT1 and PMI40 genes.
A 6.0 kb genomic DNA segment was isolated by its ability to rescue the temperature-sensitive growth defect and the hypersensitivity to sodium deoxycholate of a spontaneous vanadate-resistant mutant derived from Hansenula polymorpha DL-1. The genomic fragment contains four open reading frames homologous to the Saccharomyces cerevisiae genes YPT1 (which codes for a GTP-binding protein; 75% amino acid identity), PMI40 (encoding phosphomannose isomerase; 61% identity), YLR065c (30% identity) and CST13 (28% identity). The H. polymorpha YPT1 homologue (HpYPT1) was found to be responsible for the complementation of the temperature-sensitive phenotype and the sodium deoxycholate sensitivity of the mutant strain. Disruption of the H. polymorpha PMI40 homologue (HpPMI40) resulted in the auxotrophic requirement for D-mannose. The heterologous expressions of HpYPT1 and HpPMI40 were able to complement the temperature-sensitive phenotype of S. cerevisiae ypt1-1 mutant and the mannose auxotrophy of S. cerevisiae pmi40 null mutant, respectively, indicating that the H. polymorpha genes encode the functional homologues of S. cerevisiae YPT1 and PMI40 proteins. The nucleotide sequence has been submitted to GenBank under Accession No. AF454544.
Kim
Moo Woong
MW
Biomolecular Process Engineering Laboratory, Korea Research Institute of Bioscience and Biotechnology, Oun-dong 52, Yusong-gu, Taejon 305-600, Korea.
Agaphonov
Michael O
MO
Kim
Jeong-Yoon
JY
Rhee
Sang Ki
SK
Kang
Hyun Ah
HA
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D019204:GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005816:Genetic Complementation Test; D008358:Mannose / Q000378:metabolism; D008359:Mannose-6-Phosphate Isomerase / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010843:Pichia / Q000201:enzymology / Q000235:genetics*; D016415:Sequence Alignment; D013696:Temperature
D019204:GTP-Binding Proteins; D008359:Mannose-6-Phosphate Isomerase; D008358:Mannose