Mutation of conserved histidines alters tertiary structure and nanomechanics of consensus ankyrin repeats.

Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't

The conserved TPLH tetrapeptide motif of ankyrin repeats (ARs) plays an important role in stabilizing AR proteins, and histidine (TPLH)-to-arginine (TPLR) mutations in this motif have been associated with a hereditary human anemia, spherocytosis. Here, we used a combination of atomic force microscopy-based single-molecule force spectroscopy and molecular dynamics simulations to examine the mechanical effects of His → Arg substitutions in TPLH motifs in a model AR protein, NI6C. Our molecular dynamics results show that the mutant protein is less mechanically stable than the WT protein. Our atomic force microscopy results indicate that the mechanical energy input necessary to fully unfold the mutant protein is only half of that necessary to unfold the WT protein (53 versus 106 kcal/mol). In addition, the ability of the mutant to generate refolding forces is also reduced. Moreover, the mutant protein subjected to cyclic stretch-relax measurements displays mechanical fatigue, which is absent in the WT protein. Taken together, these results indicate that the His → Arg substitutions in TPLH motifs compromise mechanical properties of ARs and suggest that the origin of hereditary spherocytosis may be related to mechanical failure of ARs.

Full Text

Duke Authors

Cited Authors

  • Lee, W; Strümpfer, J; Bennett, V; Schulten, K; Marszalek, PE

Published Date

  • June 2012

Published In

Volume / Issue

  • 287 / 23

Start / End Page

  • 19115 - 19121

PubMed ID

  • 22514283

Electronic International Standard Serial Number (EISSN)

  • 1083-351X

Digital Object Identifier (DOI)

  • 10.1074/jbc.M112.365569

Language

  • eng

Citation Source

  • PubMed