Structural, mechanistic and functional insight into gliotoxinbis-thiomethylation inAspergillus fumigatus.
Gliotoxin is an epipolythiodioxopiperazine (ETP) class toxin, contains a disulfide bridge that mediates its toxic effects via redox cycling and is produced by the opportunistic fungal pathogenAspergillus fumigatusSelf-resistance against gliotoxin is effected by the gliotoxin oxidase GliT, and attenuation of gliotoxin biosynthesis is catalysed by gliotoxinS-methyltransferase GtmA. Here we describe the X-ray crystal structures of GtmA-apo (1.66 Å), GtmA complexed toS-adenosylhomocysteine (1.33 Å) and GtmA complexed toS-adenosylmethionine (2.28 Å), providing mechanistic insights into this important biotransformation. We further reveal that simultaneous elimination of the ability ofA. fumigatusto dissipate highly reactive dithiol gliotoxin, via deletion of GliT and GtmA, results in the most significant hypersensitivity to exogenous gliotoxin observed to date. Indeed, quantitative proteomic analysis of ΔgliT::ΔgtmAreveals an uncontrolled over-activation of thegli-cluster upon gliotoxin exposure. The data presented herein reveal, for the first time, the extreme risk associated with intracellular dithiol gliotoxin biosynthesis-in the absence of an efficient dismutation capacity. Significantly, a previously concealed protective role for GtmA and functionality of ETPbis-thiomethylation as an ancestral protection strategy against dithiol compounds is now evident.
Department of Biology, Maynooth University, Maynooth, Co. Kildare, Ireland.
Stephen K
SK
Dolan
Helmholtz Centre for Infection Research, Structure and Function of Proteins, Inhoffenstraße 7, 38124 Braunschweig, Germany.
Tobias
T
Bock
Helmholtz Centre for Infection Research, Structure and Function of Proteins, Inhoffenstraße 7, 38124 Braunschweig, Germany.
Vanessa
V
Hering
Department of Biology, Maynooth University, Maynooth, Co. Kildare, Ireland.
Rebecca A
RA
Owens
Department of Biology, Maynooth University, Maynooth, Co. Kildare, Ireland.
Gary W
GW
Jones
Helmholtz Centre for Infection Research, Structure and Function of Proteins, Inhoffenstraße 7, 38124 Braunschweig, Germany wulf.blankenfeldt@helmholtz-hzi.de.|Institute of Biochemistry, Biotechnology and Bioinformatics, Technische Universität Braunschweig, Spielmannstrasse 7, 38106 Braunschweig, Germany.
Wulf
0000-0001-9886-9668
W
Blankenfeldt
Department of Biology, Maynooth University, Maynooth, Co. Kildare, Ireland sean.doyle@nuim.ie.
Sean
0000-0003-1679-3247
S
Doyle
D001232:Aspergillus fumigatus / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D018360:Crystallography, X-Ray; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005912:Gliotoxin / Q000096:biosynthesis*; D055432:Metabolomics / Q000379:methods; D008780:Methyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D040901:Proteomics / Q000379:methods; D012435:S-Adenosylhomocysteine / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000378:metabolism; D014050:Toluene / Q000031:analogs & derivatives / Q000737:chemistry
D005656:Fungal Proteins; D014050:Toluene; D005912:Gliotoxin; D012436:S-Adenosylmethionine; D012435:S-Adenosylhomocysteine; C552832:GliT protein, Aspergillus fumigatus; D010088:Oxidoreductases; D008780:Methyltransferases; C004848:dithiol
Aspergillus; NRPS; methyltransferase; quantitative proteomics