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Quantitative Proteomic and Microarray Analysis of the Archaeon Methanosarcina acetivorans Grown with Acetate versus Methanol

Supporting Info

Lingyun Li,^†^‡ Qingbo Li,^§^‡ Lars Rohlin,^‡ UnMi Kim, Kirsty Salmon, Tomas Rejtar,^† Robert P. Gunsalus, Barry L. Karger,^† and James G. Ferry*^§

Barnett Institute and Department of Chemistry, Northeastern University, Boston, Massachusetts 02115, Department of Biochemistry and Molecular Biology, and Center for Microbial Structural Biology, 205 South Frear Laboratory, The Pennsylvania State University, University Park, Pennsylvania 16802, and Department of Microbiology, Immunology, and Molecular Genetics, and the Molecular Biology Institute, University of California, Los Angeles, California 90095

J. Proteome Res., 2007, 6 (2), pp 759–771

DOI: 10.1021/pr060383l

Publication Date (Web): December 24, 2006

†

Northeastern University.

‡

Each contributed equally.

The Pennsylvania State University.

Present address: Department of Microbiology and Immunology, University of Illinois at Chicago, Chicago, IL 60607-7173.

University of California.

To whom correspondence should be addressed. Tel.: 814/863-5721; Fax: 814/863-6217; E-mail: jgf3@psu.edu.

Abstract

Methanosarcina acetivorans strain C2A is an acetate- and methanol-utilizing methane-producing organism for which the genome, the largest yet sequenced among the Archaea, reveals extensive physiological diversity. LC linear ion trap-FTICR mass spectrometry was employed to analyze acetate- vs methanol-grown cells metabolically labeled with ¹⁴N vs ¹⁵N, respectively, to obtain quantitative protein abundance ratios. DNA microarray analyses of acetate- vs methanol-grown cells was also performed to determine gene expression ratios. The combined approaches were highly complementary, extending the physiological understanding of growth and methanogenesis. Of the 1081 proteins detected, 255 were ≥3-fold differentially abundant. DNA microarray analysis revealed 410 genes that were ≥2.5-fold differentially expressed of 1972 genes with detected expression. The ratios of differentially abundant proteins were in good agreement with expression ratios of the encoding genes. Taken together, the results suggest several novel roles for electron transport components specific to acetate-grown cells, including two flavodoxins each specific for growth on acetate or methanol. Protein abundance ratios indicated that duplicate CO dehydrogenase/acetyl-CoA complexes function in the conversion of acetate to methane. Surprisingly, the protein abundance and gene expression ratios indicated a general stress response in acetate- vs methanol-grown cells that included enzymes specific for polyphosphate accumulation and oxidative stress. The microarray analysis identified transcripts of several genes encoding regulatory proteins with identity to the PhoU, MarR, GlnK, and TetR families commonly found in the Bacteria domain. An analysis of neighboring genes suggested roles in controlling phosphate metabolism (PhoU), ammonia assimilation (GlnK), and molybdopterin cofactor biosynthesis (TetR). Finally, the proteomic and microarray results suggested roles for two-component regulatory systems specific for each growth substrate.

Keywords: quantitative proteomics • metabolic labeling • microarray • methanogenesis • acetate • methanol

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