Structural and functional studies of E. coli conjugative relaxase-helicase TraI and Arabidopsis thaliana protein arginine methyltransferase 10 (PRMT10)

Public Deposited
Analytics

Downloadable Content

Download PDF
Request Version for Screen Reader
Last Modified
  • March 22, 2019
Creator
  • Cheng, Yuan
    • Affiliation: School of Medicine, Department of Biochemistry and Biophysics
Abstract
  • TraI, a bifunctional enzyme containing relaxase and helicase activities, initiates and drives the conjugative transfer of the E. coli F plasmid. In this work, we investigated the function of the putative RecD-like domain of TraI (284-821). We established that TraI 284-821 can bind nucleotides, but only retained residual ATPase activity likely due to the loss of critical catalytic motifs. We also examined the DNA binding properties of the TraI helicase. We showed that TraI binds to single-stranded DNA (ssDNA) with a site-size of approximately 25 nucleotides and low cooperativity. A double-stranded DNA (dsDNA) binding site was identified within the N-terminal region of TraI (1-858), outside the core helicase motifs of TraI. We further characterized the impacts of ionic strength, nucleotide binding and base composition on TraI-DNA interaction. Finally, we elucidated the solution structure of TraI using small-angle x-ray scattering. Taken together, these data resulted in the assembly of a model for TraI's multi-domain helicase activity. Protein arginine methyltransferase 10 (PRMT10) regulates flowering-time in Arabidposis thaliana. Here, we present the 2.6 [angstrom] resolution crystal structure of PRMT10 that reveals significant structural features unique to PRMT10, including a long dimerization arm and distinct accessibility to the active site. Our data also showed that the N-terminal thirty residues of PRMT10 impact substrate specificity, and that PRMT10 activity was dependent on the sequences distal from the substrate methylation site. We further established that PRMT10 dimerization is required for activity and used structure-based molecular dynamics to indicate how dimerization affects functionally-essential PRMT10 domain motions. Taken together, our results provide substantial insights into the mechanism governing the unique enzymatic function and substrate specificity of PRMT10.
Date of publication
DOI
Resource type
Rights statement
  • In Copyright
Note
  • ... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Biochemistry and Biophysics.
Advisor
  • Redinbo, Matthew R.
Language

Relations

Parents:

This work has no parents.

Items

Thumbnail Title Date Uploaded Visibility Actions
file details Structural and functional studies of E. coli conjugative relaxase-helicase TraI and Arabidopsis thaliana protein arginine methyltransferase 10 (PRMT10) 2019-04-12 Public Download