Discovery and characterization of a sulfoquinovose mutarotase using kinetic analysis at equilibrium by exchange spectroscopy
AuthorAbayakoon, P; Lingford, JP; Jin, Y; Bengt, C; Davies, GJ; Yao, S; Goddard-Borger, ED; Williams, SJ
Source TitleBiochemical Journal
PublisherPORTLAND PRESS LTD
University of Melbourne Author/sGoddard-Borger, Ethan; Williams, Spencer; Yao, Shenggen; Abayakoon, Palika; Bengt, Christopher
School of Chemistry
Medical Biology (W.E.H.I.)
Document TypeJournal Article
CitationsAbayakoon, P., Lingford, J. P., Jin, Y., Bengt, C., Davies, G. J., Yao, S., Goddard-Borger, E. D. & Williams, S. J. (2018). Discovery and characterization of a sulfoquinovose mutarotase using kinetic analysis at equilibrium by exchange spectroscopy. BIOCHEMICAL JOURNAL, 475 (7), pp.1371-1383. https://doi.org/10.1042/BCJ20170947.
Access StatusOpen Access
Bacterial sulfoglycolytic pathways catabolize sulfoquinovose (SQ), or glycosides thereof, to generate a three-carbon metabolite for primary cellular metabolism and a three-carbon sulfonate that is expelled from the cell. Sulfoglycolytic operons encoding an Embden-Meyerhof-Parnas-like or Entner-Doudoroff (ED)-like pathway harbor an uncharacterized gene (yihR in Escherichia coli; PpSQ1_00415 in Pseudomonas putida) that is up-regulated in the presence of SQ, has been annotated as an aldose-1-epimerase and which may encode an SQ mutarotase. Our sequence analyses and structural modeling confirmed that these proteins possess mutarotase-like active sites with conserved catalytic residues. We overexpressed the homolog from the sulfo-ED operon of Herbaspirillum seropedicaea (HsSQM) and used it to demonstrate SQ mutarotase activity for the first time. This was accomplished using nuclear magnetic resonance exchange spectroscopy, a method that allows the chemical exchange of magnetization between the two SQ anomers at equilibrium. HsSQM also catalyzed the mutarotation of various aldohexoses with an equatorial 2-hydroxy group, including d-galactose, d-glucose, d-glucose-6-phosphate (Glc-6-P), and d-glucuronic acid, but not d-mannose. HsSQM displayed only 5-fold selectivity in terms of efficiency (kcat/KM) for SQ versus the glycolysis intermediate Glc-6-P; however, its proficiency [kuncat/(kcat/KM)] for SQ was 17 000-fold better than for Glc-6-P, revealing that HsSQM preferentially stabilizes the SQ transition state.
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