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Structural and mechanistic insights into the PAPS-independent sulfotransfer catalyzed by bacterial aryl sulfotransferase and the role of the DsbL/DsbI system in its folding

DOI: 10.1021/bi401725j DOI Help

Authors: Goran Malojcic (Harvard University) , Robin L. Owen (Diamond Light Source) , Rudi Glockshuber (ETH Zurich)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Biochemistry , VOL 53 (11) , PAGES 1870 - 1877

State: Published (Approved)
Published: March 2014

Abstract: Bacterial aryl sulfotransferases (ASSTs) catalyze sulfotransfer from a phenolic sulfate to a phenol. These enzymes are frequently found in pathogens and upregulated during infection. Their mechanistic understanding is very limited, and their natural substrates are unknown. Here, the crystal structures of Escherichia coli CFT073 ASST trapped in its presulfurylation state with model donor substrates bound in the active site are reported, which reveal the molecular interactions governing substrate recognition. Furthermore, spectroscopic titrations with donor substrates and sulfurylation kinetics of ASST illustrate that this enzyme binds substrates in a 1:1 stoichiometry and that the active sites of the ASST homooligomer act independently. Mass spectrometry and crystallographic experiments of ASST incubated with human urine demonstrate that urine contains a sulfuryl donor substrate. In addition, we examined the capability of the two paralogous dithiol oxidases present in uropathogenic E. coli CFT073, DsbA, and the ASST-specific enzyme DsbL, to introduce the single, conserved disulfide bond into ASST. We show that DsbA and DsbL introduce the disulfide bond into unfolded ASST at similar rates. Hence, a chaperone effect of DsbL, not present in DsbA, appears to be responsible for the dependence of efficient ASST folding on DsbL in vivo. The conservation of paralogous dithiol oxidases with different substrate specificities in certain bacterial strains may therefore be a consequence of the complex folding pathways of their substrate proteins.

Diamond Keywords: Enzymes; Bacteria

Subject Areas: Biology and Bio-materials, Chemistry

Facility: X06SA at Swiss Light Source

Added On: 10/06/2014 11:36


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Biochemistry Chemistry Structural biology Life Sciences & Biotech

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