Investigation of a Class of Homologous Enzymes with Sulfohydrolase Capability
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Gilmer, Caroline Rose
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Abstract
Within the metallo-ß-lactamase superfamily, an emerging and distinct class of enzymes called sulfohydrolases is being investigated. Interestingly, these diverse enzymes have been found in all kingdoms of life. To explore the differences between these types of enzymes in various organisms, we studied three homologs related to this superfamily. The first, called SdsA1 and found in the prokaryotic organism <i>Pseudomonas aeruginosa</i>, was compared to the eukaryotic enzyme Bds1 found in <i>Saccharomyces cerevisiae</i>. We found that although these two enzymes share a catalytic mechanism through nucleophilic C-attack and alkyl sulfate substrate specificity, Bds1 catalyzes the same reaction over 100 times slower. Neither Bds1 nor SdsA1 have any significant activity towards a branched primary alkyl sulfate, primary and secondary steroid sulfates, aryl sulfates, or phosphate diesters. However, SdsA1, but not Bds1, demonstrates reactivity towards a secondary alkyl sulfate. SdsA1, Bds1, and the other homologs belonging to this class that have been characterized so far exhibit a range of selectivity towards different sulfates. However another prokaryotic homolog, CddY, found in <i>Rhodococcus ruber</i> is thought to be involved in a redox reaction instead of sulfate assimilation. Currently, we are working towards expressing and purifying this homolog to test its reactivity towards a wide range of potential substrates, one of which is its proposed substrate, lauryl lactone. We also tested the reactivity of SdsA1 and Bds1 towards lauryl lactone, and our preliminary results suggest Bds1 but not SdsA1 can hydrolyze this cyclic ester. The differences in selectivity and reactivity between these homologous enzymes could mean we are gaining insight into how this class of enzymes is evolving to perform different functions.
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Biochemistry, Bds1, SdsA1, CddY, Enzyme, Kinetics, Alkyl sulfate, Aryl sulfate, Liquid chromatography-mass spectrometry, Sulfatase, Sulfohydrolase