Identification and Stereospecificity of the First Three Enzymes of 3-Dimethylsulfoniopropionate Biosynthesis in a Chlorophyte Alga1

Abstract

Many marine algae produce 3-dimethylsulfoniopropionate (DMSP), a potent osmoprotective compound whose degradation product dimethylsulfide plays a central role in the biogeochemical S cycle. Algae are known to synthesize DMSP via the four-step pathway, L-Met 3 4-methylthio-2-oxobutyrate 3 4-methylthio-2-hydroxybutyrate 3 4-dimethylsulfonio-2-hydroxy-butyrate (DM-SHB) 3 DMSP. Substrate-specific enzymes catalyzing the first three steps in this pathway were detected and partially characterized in cell-free extracts of the chlorophyte alga Enteromorpha intestinalis. The first is a 2-oxoglutarate-dependent aminotransferase, the second an NADPH-linked reductase, and the third an S-adenosylmethioninedependent methyltransferase. Sensitive radiometric assays were developed for these enzymes, and used to show that their activities are high enough to account for the estimated in vivo flux from Met to DMSP. The activities of these enzymes in other DMSP-rich chlorophyte algae were at least as high as those in E. intestinalis, but were >20-fold lower in algae without DMSP. The reductase and methyltransferase were specific for the D-enantiomer of 4-methylthio-2-hydroxybutyrate in vitro, and both the methyltransferase step and the step(s) converting DMSHB to DMSP were shown to prefer D-enantiomers in vivo. The intermediate DMSHB was shown to act as an osmoprotectant, which indicates that the first three steps of the DMSP synthesis pathway may be sufficient to confer osmotolerance.

Proteomic Analysis of a Sea-Ice Diatom: Salinity Acclimation Provides New Insight into the Dimethylsulfoniopropionate Production Pathway

“…Microalgae, such as sea-ice diatoms, are believed to utilize an alternative Met transaminase pathway. Radiolabeling studies of pathway metabolites showed algal DMSP synthesis sequentially utilizes 2-oxoglutarate-dependent aminotransferase, NADPH-linked reductase, S-adenosyl Met (SAM)-dependent methyltransferase (SAMmt), and oxidative decarboxylase (Gage et al, 1997;Summers et al, 1998). Analyzed most extensively in the green macroalgae, Ulva intestinalis, radiolabeled metabolites also supports presence of this pathway in haptophyte, diatom, and prasinophyte species.…”

“…conversion of Met to DMSP based on radiolabeling metabolite studies (Gage et al, 1997;Summers et al, 1998) would be elevated in association with salinityinduced DMSP elevation. Eighteen of the 36 proteins increased under high-salinity/DMSP conditions classified within the four enzyme classes in the proposed Met-DMSP synthesis pathway (Fig.…”

“…Although direct evidence for the involvement of Figure 6. Proteins identified in this study represent the four enzyme classes of the proposed algal Met-DMSP synthesis pathway (adapted from Gage et al, 1997;Summers et al, 1998) and active methyl cycle proteins that are important for regenerating the SAM methyl donor for the penultimate step in DMSP synthesis. Underlined proteins increased with hypersalinityinduced DMSP accumulation.…”

“…In an attempt to be more representative of natural salinity change, salinity was gradually manipulated over a 22-h period, and both pH and carbonate alkalinity were allowed to covary with salinity, as occurs within sea ice (Papadimitriou et al, 2007). We hypothesized that proteins associated with Met synthesis and from enzyme classes that drive reactions described in DMSP metabolite radiolabeling studies (Gage et al, 1997;Summers et al, 1998) will increase in abundance during hypersaline-induced DMSP accumulation. Identification of candidate proteins that may regulate DMSP synthesis will enable targeted studies on biological controls of DMSP production, which in turn can lead to improved modeling of environmental and biological feedbacks on potential DMS climate effects.…”

Sulfur isotope variability of oceanic DMSP generation and its contributions to marine biogenic sulfur emissions

“…The sulfate is subsequently transported into the chloroplasts, where it is reduced to sulfide in the presence of glutathionine and then transformed into cysteine. Cysteine is used to synthesize methionine, which is then transformed into DMSP via one of three pathways that differ among taxonomic groups of plants and algae (9)(10)(11)(12). Thus, the biosynthesis of DMSP ultimately depends on the activity of the sulfate assimilation pathway; however, little is known about how DMSP synthesis differs among algae from diverse origins, except that the whole molecule is derived from sulfur amino acids.…”