Novel triphosphate phosphohydrolase activity of Clostridium thermocellum TTM, a member of the triphosphate tunnel metalloenzyme superfamily Journal Article
| Authors: | Keppetipola, N.; Jain, R.; Shuman, S. |
| Article Title: | Novel triphosphate phosphohydrolase activity of Clostridium thermocellum TTM, a member of the triphosphate tunnel metalloenzyme superfamily |
| Abstract: | Triphosphate tunnel metalloenzymes (TTMs) are a newly recognized superfamily of phosphotransferases defined by a unique active site residing within an eight-stranded β barrel. The prototypical members are the eukaryal metal-dependent RNA triphosphatases, which catalyze the initial step in mRNA capping. Little is known about the activities and substrate specificities of the scores of TTM homologs present in bacterial and archaeal proteomes, nearly all of which are annotated as adenylate cyclases. Here we have conducted a biochemical and structure-function analysis of a TTM protein (CthTTM) from the bacterium Clostridium thermocellum. CthTTM is a metal-dependent tripolyphosphatase and nucleoside triphosphatase; it is not an adenylate cyclase. We have identified 11 conserved amino acids in the tunnel that are critical for tripolyphosphatase and ATPase activity. The most salient findings are that (i) CthTTM is 150-fold more active in cleaving tripolyphosphate than ATP and (ii) the substrate specificity of CthTTM can be transformed by a single mutation (K8A) that abolishes tripolyphosphatase activity while strongly stimulating ATP hydrolysis. Our results underscore the plasticity of CthTTM substrate choice and suggest how novel specificities within the TTM superfamily might evolve through changes in the residues that line the tunnel walls. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc. |
| Keywords: | controlled study; unclassified drug; mutation; nonhuman; molecular genetics; protein function; metabolism; phosphatase; protein degradation; enzymology; nucleoside triphosphatase; enzyme activity; structure activity relation; structure-activity relationship; bacteria (microorganisms); physiology; time; time factors; rna; chemistry; amino acid sequence; molecular sequence data; eukaryota; recombinant proteins; substrate specificity; recombinant protein; binding site; models, molecular; binding sites; chemical structure; alanine; amino acids; adenosine triphosphate; molecular biology; protein family; enzyme specificity; enzyme structure; biochemistry; phosphates; hydrolysis; phosphoric monoester hydrolases; enzymes; archaea; metal; metals; adenylate cyclase; clostridium thermocellum; metalloenzyme; triphosphate tunnel metalloenzyme; nucleoside-triphosphatase; cleaving tripolyphosphate; triphosphate tunnel metalloenzymes (ttm); tripolyphosphatase activity |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 282 |
| Issue: | 16 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2007-04-20 |
| Start Page: | 11941 |
| End Page: | 11949 |
| Language: | English |
| DOI: | 10.1074/jbc.M611328200 |
| PUBMED: | 17303560 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | --- - "Cited By (since 1996): 6" - "Export Date: 17 November 2011" - "CODEN: JBCHA" - "Source: Scopus" |
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