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Microbial pathway for anaerobic 5′-methylthioadenosine metabolism coupled to ethylene formation

  1. F. Robert Tabitaa,1
  1. aDepartment of Microbiology, The Ohio State University, Columbus, OH 43210
  1. Edited by Bob B. Buchanan, University of California, Berkeley, CA, and approved October 20, 2017 (received for review June 29, 2017)

  1. Fig. 2.

    Organization of known and putative MTA metabolism gene clusters. (A) Gene clusters from representative organisms containing mtnP or mtnK, mtnA, and ald2 aligned to R. rubrum as part of a putative anaerobic MSP (SI Appendix, Table S5). Note that certain Bacillus sp. (e.g., B. cereus, B. anthracis, B. thuringiensis) possess an entirely separate gene cluster for the universal MSP (Fig. 1, black and blue arrows). (B) MTA-isoprenoid shunt-specific gene cluster from R. rubrum and R. palustris.

  2. Fig. 3.

    Ethylene production in the presence of various sulfur sources. (A and B) Total ethylene produced [micromoles of ethylene per liter culture per optical density measured at 660 nm (μmol/L/OD660 nm)] by R. rubrum and R. palustris, respectively, when grown to stationary phase on l-methionine (Met), MTA, or 2-(methylthio)ethanol (MT-EtOH) supplied at 1 mM. (C and D) R. rubrum ethylene induction after switching to anaerobic growth on 2-(methylthio)ethanol. Cultures were initially grown aerobically on 1 mM ammonium sulfate (triangles), anaerobically on 1 mM ammonium sulfate (circles), or anaerobically on 1 mM 2-(methylthio)ethanol (squares) before being washed into anaerobic media containing 1 mM 2-(methylthio)ethanol without or with 15 μg/mL chloramphenicol. Error bars are SDs from n = 3 independent induction experiments. Dashed lines are fits of data to a sigmoidal-logistic model to determine onset of ethylene production given by when ethylene exceeds the 0.01 μmol/L/OD660?nm detection limit.

  3. Fig. 4.

    Identification of reaction catalyzed by Ald2. (A) GC traces identifying specific conversion of 5-(methylthio)ribulose-1-P to 2-(methylthio)acetaldehdye catalyzed by purified R. rubrum Ald2 protein. Conversion of 5-(methylthio)ribose-1-P to 5-(methylthio)ribulose-1-P is catalyzed by purified R. rubrum 5-(methylthio)ribulose-1-P isomerase protein (MtnA). (B) GC traces identifying specific conversion of 2-(methylthio)acetaldehyde to 2-(methylthio)ethanol by yeast ADH. (C and D) [methyl-14C]-Metabolites produced by R. rubrum strains Δrlp1 and Δrlp1/Δald2, respectively, at the indicated time (h) after feeding [methyl-14C]MTA. (E) [methyl-14C]2-(Methylthio)ethanol observed in the indicated R. rubrum strain at the indicated time after feeding [methyl-14C]MTA. (F) Ethylene stoichiometry measurements from 2-(methylthio)ethanol. Total ethylene produced (in nanomoles) from the indicated amount of 2-(methylthio)ethanol (in nanomoles) fed to R. rubrum (circles) or R. palustris (squares). Error bars are SD from three independent feedings. Data were fit to a linear regression. Compounds: (i) 5-(methylthio)ribose-1-P (MTR-1P), (ii) 5-(methylthio)ribulose-1-P (MTRu-1P), (iii) 2-(methylthio)acetaldehyde (MT-adh), (iv) 5-(methylthio)ethanol (MT-EtOH), (v) 5-(methylthio)ribose, (vi) 5-(methylthio)ribulose, (vii) unknown not involved in anaerobic ethylene-forming MSP, and (viii) unknown contaminant present in commercial [methyl-14C]SAM.

  4. Fig. 5.

    Ethylene-forming pathways. (A) Ethylene pathway involving ethylene-forming enzyme (EFE) present in some bacterial and fungi. (B) Nonenzymatic ethylene pathways involving flavin or hydroxyl radial attack of KMTB or methional and methionol (not shown). Photooxidation of flavin is the only previously known path to ethylene not requiring oxygen. (C) Plant ethylene biosynthesis pathway via ACC metabolism from SAM. (D) Anaerobic ethylene-forming MSP from R. rubrum and R. palustris.

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