| First Author | Mukai M | Year | 2009 |
| Journal | Biochemistry | Volume | 48 |
| Issue | 5 | Pages | 1084-93 |
| PubMed ID | 19152315 | Mgi Jnum | J:149071 |
| Mgi Id | MGI:3847579 | Doi | 10.1021/bi802047y |
| Citation | Mukai M, et al. (2009) Recombinant mammalian tubulin polyglutamylase TTLL7 performs both initiation and elongation of polyglutamylation on beta-tubulin through a random sequential pathway. Biochemistry 48(5):1084-93 |
| abstractText | Tubulins undergo unique post-translational modifications, such as tyrosination, polyglutamylation, and polyglycylation. These modifications are performed by members of a protein family, the tubulin tyrosine ligase (TTL)-like (TTLL) family, which is characterized by the presence of a highly conserved TTL domain. We and others have recently identified tubulin polyglutamylases in the TTLL family [Janke, C., et al. (2005) Science 308, 1758-1762; Ikegami, K., et al. (2006) J. Biol. Chem. 281, 30707-30716; van Dijk, J., et al. (2007) Mol. Cell 26, 437-448]. Previously, we identified TTLL7 as a beta-tubulin-selective polyglutamylase. However, there is controversy over whether TTLL7 functions as an initiase, elongase, or both in polyglutamylation. In this report, we investigate the polyglutamylation reaction by TTLL7 by employing a recombinant enzyme and in vitro reaction. Two-dimensional electrophoresis and tandem mass spectrometry showed that TTLL7 performed both the initiation and elongation of polyglutamylation on beta-tubulin. Recombinant TTLL7 performed with a maximal and specific activity to polymerized tubulin at a neutral pH and a lower salt concentration. The initial rate and inhibitor analyses revealed that the mechanism of binding of three substrates, glutamate, ATP, and tubulin, to the enzyme was a random sequential pathway. Our findings provide evidence that mammalian TTLL7 performs both initiation and elongation in the polyglutamylation reaction on beta-tubulin through a random sequential pathway. |
