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Title: A hinge migration mechanism unlocks the evolution of green-to-red photoconversion in GFP-like proteins

In proteins, functional divergence involves mutations that modify structure and dynamics. In this paper, we provide experimental evidence for an evolutionary mechanism driven solely by long-range dynamic motions without significant backbone adjustments, catalytic group rearrangements, or changes in subunit assembly. Crystallographic structures were determined for several reconstructed ancestral proteins belonging to a GFP class frequently employed in superresolution microscopy. Their chain flexibility was analyzed using molecular dynamics and perturbation response scanning. The green-to-red photoconvertible phenotype appears to have arisen from a common green ancestor by migration of a knob-like anchoring region away from the active site diagonally across the β barrel fold. The allosterically coupled mutational sites provide active site conformational mobility via epistasis. We propose that light-induced chromophore twisting is enhanced in a reverse-protonated subpopulation, activating internal acid-base chemistry and backbone cleavage to enlarge the chromophore. Finally, dynamics-driven hinge migration may represent a more general platform for the evolution of novel enzyme activities.
 [1] ;  [2] ;  [3] ;  [2] ;  [2] ;  [2] ;  [2] ;  [3] ;  [2] ;  [2]
  1. Arizona State Univ., Tempe, AZ (United States); Univ. of Michigan Medical School, Ann Arbor, MI (United States)
  2. Arizona State Univ., Tempe, AZ (United States)
  3. The Univ. of Texas, Austin, TX (United States)
Publication Date:
OSTI Identifier:
Grant/Contract Number:
AC02-05CH11231; AC02-06CH11357
Published Article
Journal Name:
Additional Journal Information:
Journal Volume: 23; Journal Issue: 1; Journal ID: ISSN 0969-2126
Research Org:
Arizona State Univ., Tempe, AZ (United States)
Sponsoring Org:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
Country of Publication:
United States
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; green fluorescent protein; red fluorescent protein; superresolution microscopy; Kaede; photoconversion; photochromism; ancestral gene reconstruction; molecular evolution; protein evolution; protein dynamics; chain flexibility; catalytic base activation; reverse protonation of ionic bonds; principle of kinetic equivalence; allosteric communication