Molecular and genetic analysis of the Cryptococcus neoformans MET3 gene and a met3 mutant a aThe GenBank accession numbers for the sequences reported in this paper are AY035556 and AF489498.

Abstract

The Cryptococcus neoformans MET3 cDNA (encoding ATP sulfurylase) was cloned by complementation of the corresponding met3 mutation in Saccharomyces cerevisiae. Sequence analysis showed high similarity between the deduced amino acid sequence of the C. neoformans Met3p and other fungal ATP sulfurylases. A C. neoformans met3 mutant was made by targeted insertional mutagenesis, which had the expected auxotrophic phenotype, and reconstituted the met3 mutant to Met M . In vitro, the C. neoformans met3 mutant had a substantial defect in melanin formation, significantly reduced growth rate, and greatly increased thermotolerance. In the murine inhalation infection model, the met3 mutant was avirulent and was deficient in its ability to survive in mice. It is concluded that, in contrast to the yeast form of Histoplasma capsulatum, in C. neoformans the sulfate-assimilation arm of the methionine biosynthetic pathway plays an important role in vitro, even in the presence of abundant exogenous methionine, and is critical for virulence, and indeed for survival, in vivo.

Cryptococcus neoformans Ilv2p confers resistance to sulfometuron methyl and is required for survival at 37 °C and in vivo

“…Amino acid biosynthetic pathways are interesting from the perspective of potential antifungal drug targets as (1) various fungal auxotrophs are reduced in virulence (see, for example, Goldstein & McCusker, 2001;Manning et al, 1984;Namiki et al, 2001;Yang et al, 2002); (2) these pathways are conserved throughout fungi; (3) many of these pathways do not exist in humans; and (4) various amino acid biosynthetic inhibitors have already been identified for use as herbicides. The branched-chain amino acids (isoleucine, valine and leucine) are examples of amino acids that can be synthesized by fungi, but which are essential in mammals.…”

Cystathionine β-Lyase Is Important for Virulence of Salmonella enterica Serovar Typhimurium

“…The biosynthesis of Met is therefore of vital importance to microbial growth. This has been validated by the fact that several natural products, including 2-amino-5-hydroxy-4-oxopentanoic acid (8), azoxybacilin (1), and rhizocticin (10), target important enzymes for Met biosynthesis and have antimicrobial properties and that the disruption of genes such as STR3, which is important in the fungal sulfur assimilation pathway, results in avirulence in a Cryptococcus neoformans infection model (17).…”

β-Lactone natural products and derivatives inactivate homoserine transacetylase, a target for antimicrobial agents

“…[1][2][3][4] This pathway uses several enzymes to produce the essential amino acids methionine (Met), Thr and Ile, which are not produced in mammals. 5 The absence of this pathway in mammals makes it an attractive target for antimicrobial drug discovery.…”

“…3,[12][13][14] Moreover, gene disruption methods have identified the importance of these enzymes in pathogenesis in several animal models of virulence. 1,2,4,[15][16][17] One key enzyme in the Asp pathway is HTA, which catalyzes the transfer of an acetyl group from acetyl-CoA to the hydroxyl group of homoserine ( Figure 1). 18 This enzyme catalyzes the first committed step in the biosynthesis of Met from aspartic acid.…”