HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Yamada-Okabe, T. Articles by Yamada-Okabe, H. Search for Related Content PubMed PubMed Citation Articles by Yamada-Okabe, T. Articles by Yamada-Okabe, H. Agricola Articles by Yamada-Okabe, T. Articles by Yamada-Okabe, H.

The Candida albicans gene for mRNA 5'-cap methyltransferase: identification of additional residues essential for catalysis

Department of Hygiene, School of Medicine, Yokohama City University, 3-9 Fukuura, Kanazawa, Yokohama 236-0004, Japan¹
Department of Mycology, Nippon Roche Research Center, 200 Kajiwara, Kamakura, Kanagawa 247-8530, Japan²

Author for correspondence: Hisafumi Yamada-Okabe. Tel: +81 467 45 4382. Fax: +81 467 46 5320. e-mail: hisafumi.okabe{at}roche.com

The 5'-cap structure of eukaryotic mRNA is methylated at the terminal guanosine by RNA (guanine-N⁷-)-methyltransferase (cap MTase). Saccharomyces cerevisiae ABD1 (ScABD1) and human hMet (also called CMT1) genes are responsible for this enzyme. The ABD1 homologue was cloned from the pathogenic fungus Candida albicans and named C. albicans ABD1 (CaABD1). When expressed as a fusion with glutathione S-transferase (GST), CaAbd1p displayed cap MTase activity in vitro and rescued S. cerevisiae abd1 null mutants, indicating that CaABD1 specifies an active cap MTase. Although the human cap MTase binds to the human capping enzyme (Hce1p), which possesses both mRNA guanylyltransferase (mRNA GTase) and mRNA 5'-triphosphatase (mRNA TPase) activities, yeast two-hybrid analysis demonstrated that in yeast neither mRNA GTase nor mRNA TPase physically interacted with the Abd1 protein. Comparison of the amino acid sequences of known and putative cap MTases revealed a highly conserved amino acid sequence motif, Phe/Val-Leu-Asp/Glu-Leu/Met-Xaa-Cys-Gly-Lys-Gly-Gly-Asp-Leu-Xaa-Lys, which encompasses the sequence motif characteristic of divergent methyltransferases. Mutations in CaAbd1p of leucine at the second and the twelfth positions (so far uncharacterized) to alanine severely diminished the enzyme activity and the functionality in vivo, whereas those of leucine at the fourth, cysteine at the sixth, lysine at the eighth, and glycine at the tenth positions did not. Furthermore, valine substitution for the twelfth, but not for the second, leucine in that motif abolished the activity and functionality of CaAbd1p. Thus, it appears that leucine at the second and the twelfth positions in the motif, together with a previously identified acidic residue in the third, glycine at the sixth and glutamic acid at the eleventh positions, play important roles in the catalysis, and that side chain length is crucial for the activity at the twelfth position in the motif.

Keywords: mRNA 5'-capping, mRNA cap methyltransferase, human, cDNA, Candida albicans

Abbreviations: cap MTase, RNA (guanine-N⁷-)-methyltransferase; 5-FOA, 5-fluoroorotic acid; GST, glutathione S-transferase; mRNA GTase, mRNA guanylyltransferase; mRNA TPase, mRNA triphosphatase; SAM, S-adenosyl-L-methionine

The GenBank accession numbers for the nucleotide sequence of CaABD1 and hMet are AB020965 and AB020966, respectively.

This article has been cited by other articles:

				S. Zheng, S. Hausmann, Q. Liu, A. Ghosh, B. Schwer, C. D. Lima, and S. Shuman Mutational Analysis of Encephalitozoon cuniculi mRNA Cap (Guanine-N7) Methyltransferase, Structure of the Enzyme Bound to Sinefungin, and Evidence That Cap Methyltransferase Is the Target of Sinefungin's Antifungal Activity J. Biol. Chem., November 24, 2006; 281(47): 35904 - 35913. [Abstract] [Full Text] [PDF]

				V. Z. Grdzelishvili, S. Smallwood, D. Tower, R. L. Hall, D. M. Hunt, and S. A. Moyer A Single Amino Acid Change in the L-Polymerase Protein of Vesicular Stomatitis Virus Completely Abolishes Viral mRNA Cap Methylation J. Virol., June 15, 2005; 79(12): 7327 - 7337. [Abstract] [Full Text] [PDF]

				G. L. Chrebet, D. Wisniewski, A. L. Perkins, Q. Deng, M. B. Kurtz, A. Marcy, and S. A. Parent Cell-Based Assays to Detect Inhibitors of Fungal mRNA Capping Enzymes and Characterization of Sinefungin as a Cap Methyltransferase Inhibitor J Biomol Screen, June 1, 2005; 10(4): 355 - 364. [Abstract] [PDF]

				S. Hausmann, S. Zheng, C. Fabrega, S. W. Schneller, C. D. Lima, and S. Shuman Encephalitozoon cuniculi mRNA Cap (Guanine N-7) Methyltransferase: METHYL ACCEPTOR SPECIFICITY, INHIBITION BY S-ADENOSYLMETHIONINE ANALOGS, AND STRUCTURE-GUIDED MUTATIONAL ANALYSIS J. Biol. Chem., May 27, 2005; 280(21): 20404 - 20412. [Abstract] [Full Text] [PDF]

				Y. Takase, T. Takagi, P. B. Komarnitsky, and S. Buratowski The Essential Interaction between Yeast mRNA Capping Enzyme Subunits Is Not Required for Triphosphatase Function In Vivo Mol. Cell. Biol., December 15, 2000; 20(24): 9307 - 9316. [Abstract] [Full Text]

				B. Schwer, K. Lehman, N. Saha, and S. Shuman Characterization of the mRNA Capping Apparatus of Candida albicans J. Biol. Chem., January 12, 2001; 276(3): 1857 - 1864. [Abstract] [Full Text] [PDF]