Structural and kinetic insights into the stereospecific oxidation of 2,3-dihydroxypropanesulfonate by DHPS-3-dehydrogenase (HpsN) from Cupriavidus pinatubonensis

30 November 2023, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


2,3-Dihydroxypropanesulfonate (DHPS) and sulfolactate (SL) are environmentally important organosulfur compounds that play key roles as metabolic currencies in the sulfur cycle. Despite their prevalence, the pathways governing DHPS and SL production remain poorly understood. In this study, we explore the DHPS-3-dehydrogenase CpHpsN from Cupriavidus pinatubonensis, a bacterium capable of utilizing DHPS as a sole carbon source. Kinetic analysis of CpHpsN reveals a strict preference for R-DHPS, catalyzing its 4-electron oxidation to R-SL, with high specificity for NAD+ over NADP+. The 3D structure of CpHpsN, elucidated through X-ray crystallography, unveils a fold akin to E. coli histidinol dehydrogenase (HisD), albeit with key differences around the Zn2+ centre that provide the structural basis for substrate specificity. These differences arise from a change in position of a protein loop that is a key sequence that distinguishes HpsN and HisD homologues. Site-directed mutagenesis pinpoint Glu318, His319, and Asp352 as active site residues important for the catalytic activity of CpHpsN. Taxonomic and pathway distribution analysis reveals the prevalence of HpsN within different pathways of DHPS catabolism and across bacterial classes including Alpha-, Beta-, Gamma-, and Deltaproteobacteria and Desulfobacteria, emphasizing its importance in the biogeochemical sulfur cycle.


enzyme catalysis
protein structure
biogeochemical sulfur cycle
microbial metabolism

Supplementary materials

Supplementary Information
Experimental procedures, Supplementary figures and table


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