Crystal structure and biochemical characterization of a Mycobacterium smegmatis AAA-type nucleoside triphosphatase phosphohydrolase (Msm0858) Journal Article
| Authors: | Unciuleac, M. C.; Smith, P. C.; Shuman, S. |
| Article Title: | Crystal structure and biochemical characterization of a Mycobacterium smegmatis AAA-type nucleoside triphosphatase phosphohydrolase (Msm0858) |
| Abstract: | AAA proteins (ATPases associated with various cellular activities) use the energy of ATP hydrolysis to drive conformational changes in diverse macromolecular targets. Here, we report the biochemical characterization and 2.5-Å crystal structure of a Mycobacterium smegmatis AAA protein Msm0858, the ortholog of Mycobacterium tuberculosis Rv0435c. Msm0858 is a magnesium- dependent ATPase and is active with all nucleoside triphosphates (NTPs) and deoxynucleoside triphosphates (dNTPs) as substrates. The Msm0858 structure comprises (i) an N-terminal domain (amino acids [aa] 17 to 201) composed of two β-barrel modules and (ii) two AAA domains, D1 (aa 212 to 473) and D2 (aa 476 to 744), each of which has ADP in the active site. Msm0858-ADP is a monomer in solution and in crystallized form. Msm0858 domains are structurally homologous to the corresponding modules of mammalian p97. However, the position of the N-domain modules relative to the AAA domains in the Msm0858-ADP tertiary structure is different and would impede the formation of a p97-like hexameric quaternary structure. Mutational analysis of the A-box and B-box motifs indicated that the D1 and D2 AAA domains are both capable of ATP hydrolysis. Simultaneous mutations of the D1 and D2 active-site motifs were required to abolish ATPase activity. ATPase activity was effaced by mutation of the putative D2 arginine finger, suggesting that Msm0858 might oligomerize during the ATPase reaction cycle. A truncated variant Msm0858 (aa 212 to 745) that lacks the N domain was characterized as a catalytically active homodimer. © 2016, American Society for Microbiology. |
| Journal Title: | Journal of Bacteriology |
| Volume: | 198 |
| Issue: | 10 |
| ISSN: | 0021-9193 |
| Publisher: | American Society for Microbiology |
| Date Published: | 2016-05-01 |
| Start Page: | 1521 |
| End Page: | 1533 |
| Language: | English |
| DOI: | 10.1128/jb.00905-15 |
| PROVIDER: | scopus |
| PMCID: | PMC4859601 |
| PUBMED: | 26953339 |
| DOI/URL: | |
| Notes: | Article -- Export Date: 2 August 2016 -- Source: Scopus |
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