PMID: 16801438 A Phe389Leu substitution in ergA confers terbinafine resistance in Aspergillus fumigatus

MPiovesana commented 1 year ago

Paper curated by @MPiovesana https://canto.phi-base.org/curs/6b98cb8ecdf4aa5f See latest comment for updated info on genotype creation!

MPiovesana commented 1 year ago

Uniprot ID: The authors provide three GenBank accession numbers ('A nucleotide sequence with 73% sequence similarity to ErgA was obtained from the A. fumigatus genome database (GenBank accession no. EAL91820, AY619002.1, and AY532916'). The first one seems to be the best option as it is the curated entry for the squalene epoxidase of A. fumigatus erg1. The second accession number does not return any entries on Uniprot, and the third one returns a non-curated entry. Thus, I opted for Uniprot ID associated with GenBank entry EAL91820, which is E9R5G2.

AC: search uniprot with 'EAL91820' gives swiss-prot reviewed entry https://www.uniprot.org/uniprotkb/E9R5G2/entry

MPiovesana commented 1 year ago

Question: Authors mention that "Plasmid pRG47 also contains a silent SacII site mutation within the ergA gene fragment to facilitate rapid identification of transformants"; the site of this silent mutation is not provided. Is it necessary to mention this in the comment section of the genotype annotation, perhaps?

AC: for now I have not added this as we don't have the site of this silent mutation.

MPiovesana commented 1 year ago

Curation complete pending review.

MPiovesana commented 11 months ago

UPDATED After revisiting this session and reading the paper again, I believe allele type transformant is not the best option to curate this genotype. Authors clone the wild type ergA gene and introduce a point mutation by in vitro mutagenesis. The mutant allele is then introduced into a wild type strains, and the resulting genotypes display chemical resistance phenotypes. Despite the authors not mentioning which wild type strain was used for the amplification of the wt ergA gene, I think it can be assumed from the text that it was the same strain used as a recipient for the transformation, and therefore, I think allele type amino acid substitution would be the best option here. Thus, the genotype below was created:

ErgA-F389L(aaF389L)[Not assayed]

As authors state 'DNA analysis revealed the ergA gene to be ectopically integrated into the TRBR-Ec (TRB-resistant ectopic) strain, indicating that expression of the ectopic mutant gene was dominant over the wild-type allele' I selected ectopic for expression level. The presence of the endogenous wt ErgA gene was recorded in background.