Atomic-resolution structure of the CAP-Gly domain of dynactin on polymeric microtubules determined by magic angle spinning NMR spectroscopy

Proc Natl Acad Sci U S A. 2015 Nov 24;112(47):14611-6. doi: 10.1073/pnas.1509852112. Epub 2015 Nov 10.

Abstract

Microtubules and their associated proteins perform a broad array of essential physiological functions, including mitosis, polarization and differentiation, cell migration, and vesicle and organelle transport. As such, they have been extensively studied at multiple levels of resolution (e.g., from structural biology to cell biology). Despite these efforts, there remain significant gaps in our knowledge concerning how microtubule-binding proteins bind to microtubules, how dynamics connect different conformational states, and how these interactions and dynamics affect cellular processes. Structures of microtubule-associated proteins assembled on polymeric microtubules are not known at atomic resolution. Here, we report a structure of the cytoskeleton-associated protein glycine-rich (CAP-Gly) domain of dynactin motor on polymeric microtubules, solved by magic angle spinning NMR spectroscopy. We present the intermolecular interface of CAP-Gly with microtubules, derived by recording direct dipolar contacts between CAP-Gly and tubulin using double rotational echo double resonance (dREDOR)-filtered experiments. Our results indicate that the structure adopted by CAP-Gly varies, particularly around its loop regions, permitting its interaction with multiple binding partners and with the microtubules. To our knowledge, this study reports the first atomic-resolution structure of a microtubule-associated protein on polymeric microtubules. Our approach lays the foundation for atomic-resolution structural analysis of other microtubule-associated motors.

Keywords: dynactin's CAP-Gly domain; intermolecular interface determination; magic angle spinning NMR; microtubules; structure determination.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Calibration
  • Dynactin Complex
  • Glycine / chemistry*
  • Magnetic Resonance Spectroscopy / methods*
  • Microtubule-Associated Proteins / chemistry*
  • Microtubules / metabolism*
  • Models, Molecular
  • Molecular Sequence Data
  • Polymerization*
  • Protein Structure, Tertiary

Substances

  • Dynactin Complex
  • Microtubule-Associated Proteins
  • Glycine

Associated data

  • PDB/2MPX