Characterization, heterologous expression and functional analysis of mevalonate diphosphate decarboxylase gene (MVD) of Candida albicans.
Mevalonate diphosphate decarboxylase (MVD) catalyzes the conversion of mevalonate diphosphate to isopentenyl diphosphate, a key building block for a large family of functionally important biomolecules. We have cloned the gene encoding MVD from Candida albicans, and report the first characterization of such a gene from an opportunistic fungal pathogen. Sequence analysis revealed that the MVD comprises 362 amino acid residues with a predicted molecular weight of 39.5 kDa, sharing minimal identity with the human analogue. Analysis of the genomic sequence indicated that the coding frame is interrupted by a small intron of 51 bp. Southern analysis of genomic DNA demonstrated that MVD is a single-copy gene. Furthermore, Southern analysis of electrophoretic karyotypes of C. albicans obtained by pulsed-field gel electrophoresis showed that MVD is located on chromosome 1. Northern analysis revealed that the level of MVD expression is affected by (1) the carbon source in the growth medium, (2) the growth phase, and (3) the growth form of the fungus (yeast-like or hyphal). To demonstrate the biological function of C. albicans MVD, complementation experiments were carried out with an S. cerevisiae strain (erg19(ts)) that is temperature-sensitive for MVD activity. A single copy of the C. albicans MVD gene, under the control of the NOP1 promoter, was able fully to complement the erg19(ts) phenotype, and expression of the epitope-tagged C. albicans MVD was detectable by Western analysis. Furthermore, the low degree of sequence identity between C. albicans MVD and its human analogue raises the possibility that fungal-specific inhibitors can be developed for the enzyme. Thus, C. albicans MVD appears to be an interesting candidate that could be targeted for the development of anti-fungal agents.
Dassanayake
R S
RS
Department of Oral Biosciences, Faculty of Dentistry, University of Hong Kong, Prince Philip Dental Hospital, Hong Kong SAR, China.
Cao
L
L
Samaranayake
L P
LP
Berges
T
T
D000595:Amino Acid Sequence; D001483:Base Sequence; D002176:Candida albicans / Q000201:enzymology* / Q000235:genetics; D002262:Carboxy-Lyases / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D010802:Phylogeny; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D017422:Sequence Analysis, DNA
D002262:Carboxy-Lyases; C022503:pyrophosphomevalonate decarboxylase