期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 110, 期 21, 页码 8519-8524出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1302417110
关键词
iron-sulfur cluster fold; radical SAM dehydrogenase
资金
- National Institutes of Health (NIH) [GM-63847, GM-103268]
- National Science Foundation [MCB-0543833]
- National Center for Research Resources at NIH [RR-15301]
- US DOE [DE-AC02-06CH11357]
- Office of Biological and Environmental Research
- Office of Basic Energy Sciences of the US DOE
- National Center for Research Resources [P41RR012408]
- National Institute of General Medical Sciences of NIH [P41GM103473]
Arylsulfatases require a maturating enzyme to perform a co- or posttranslational modification to form a catalytically essential formylglycine (FGly) residue. In organisms that live aerobically, molecular oxygen is used enzymatically to oxidize cysteine to FGly. Under anaerobic conditions, S-adenosylmethionine (AdoMet) radical chemistry is used. Here we present the structures of an anaerobic sulfatase maturating enzyme (anSME), both with and without peptidyl-substrates, at 1.6-1.8 angstrom resolution. We find that anSMEs differ from their aerobic counterparts in using backbone-based hydrogen-bonding patterns to interact with their peptidyl-substrates, leading to decreased sequence specificity. These anSME structures from Clostridium perfringens are also the first of an AdoMet radical enzyme that performs dehydrogenase chemistry. Together with accompanying mutagenesis data, a mechanistic proposal is put forth for how AdoMet radical chemistry is coopted to perform a dehydrogenation reaction. In the oxidation of cysteine or serine to FGly by anSME, we identify D277 and an auxiliary [4Fe-4S] cluster as the likely acceptor of the final proton and electron, respectively. D277 and both auxiliary clusters are housed in a cysteinerich C-terminal domain, termed SPASM domain, that contains homology to similar to 1,400 other unique AdoMet radical enzymes proposed to use [4Fe-4S] clusters to ligate peptidyl-substrates for subsequent modification. In contrast to this proposal, we find that neither auxiliary cluster in anSME bind substrate, and both are fully ligated by cysteine residues. Instead, our structural data suggest that the placement of these auxiliary clusters creates a conduit for electrons to travel from the buried substrate to the protein surface.
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