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Dynamics of Aromatic Side Chains in the Active Site of FKBP12

Weininger, Ulrich LU ; Modig, Kristofer LU ; Geitner, Anne Juliane; Schmidpeter, Philipp A.M.; Koch, Johanna R. and Akke, Mikael LU (2017) In Biochemistry 56(1). p.334-343
Abstract

FKBP12, a small human enzyme, aids protein folding by catalyzing cis-trans isomerization of peptidyl-prolyl bonds, and is involved in cell signaling pathways, calcium regulation, and the immune response. The underlying molecular mechanisms are not fully understood, but it is well-known that aromatic residues in the active site and neighboring loops are important for substrate binding and catalysis. Here we report micro- to millisecond exchange dynamics of aromatic side chains in the active site region of ligand-free FKBP12, involving a minor state population of 0.5% and an exchange rate of 3600 s-1, similar to previous results for the backbone and methyl-bearing side chains. The exchange process involves tautomerization of H87. In the... (More)

FKBP12, a small human enzyme, aids protein folding by catalyzing cis-trans isomerization of peptidyl-prolyl bonds, and is involved in cell signaling pathways, calcium regulation, and the immune response. The underlying molecular mechanisms are not fully understood, but it is well-known that aromatic residues in the active site and neighboring loops are important for substrate binding and catalysis. Here we report micro- to millisecond exchange dynamics of aromatic side chains in the active site region of ligand-free FKBP12, involving a minor state population of 0.5% and an exchange rate of 3600 s-1, similar to previous results for the backbone and methyl-bearing side chains. The exchange process involves tautomerization of H87. In the major state H87 is highly flexible and occupies the common HNε2 tautomer, while in the minor state it occupies the rare HNδ1 tautomer, which typically requires stabilization by specific interactions, such as hydrogen bonds. This finding suggests that the exchange process is coupled to a rearrangement of the hydrogen bond network around H87. Upon addition of the active-site inhibitor FK506 the exchange of all aromatic residues is quenched, with exception of H87. The H87 resonances are broadened beyond detection, suggesting that interconversion between tautomers prevail in the FK506-bound state. While key active-site residues undergo conformational exchange in the apo state, the exchange rate is considerably faster than the catalytic turnover, as determined herein by Michaelis-Menten type analysis of NMR line shapes and chemical shifts. We discuss alternative interpretations of this observation in terms of FKBP12 function.

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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Biochemistry
volume
56
issue
1
pages
10 pages
publisher
The American Chemical Society
external identifiers
  • scopus:85020323996
  • wos:000391898400033
ISSN
0006-2960
DOI
10.1021/acs.biochem.6b01157
language
English
LU publication?
yes
id
9597019c-193d-49c5-b2bd-404ae8e4ca14
date added to LUP
2017-06-29 11:21:39
date last changed
2017-09-18 11:37:00
@article{9597019c-193d-49c5-b2bd-404ae8e4ca14,
  abstract     = {<p>FKBP12, a small human enzyme, aids protein folding by catalyzing cis-trans isomerization of peptidyl-prolyl bonds, and is involved in cell signaling pathways, calcium regulation, and the immune response. The underlying molecular mechanisms are not fully understood, but it is well-known that aromatic residues in the active site and neighboring loops are important for substrate binding and catalysis. Here we report micro- to millisecond exchange dynamics of aromatic side chains in the active site region of ligand-free FKBP12, involving a minor state population of 0.5% and an exchange rate of 3600 s-1, similar to previous results for the backbone and methyl-bearing side chains. The exchange process involves tautomerization of H87. In the major state H87 is highly flexible and occupies the common HNε2 tautomer, while in the minor state it occupies the rare HNδ1 tautomer, which typically requires stabilization by specific interactions, such as hydrogen bonds. This finding suggests that the exchange process is coupled to a rearrangement of the hydrogen bond network around H87. Upon addition of the active-site inhibitor FK506 the exchange of all aromatic residues is quenched, with exception of H87. The H87 resonances are broadened beyond detection, suggesting that interconversion between tautomers prevail in the FK506-bound state. While key active-site residues undergo conformational exchange in the apo state, the exchange rate is considerably faster than the catalytic turnover, as determined herein by Michaelis-Menten type analysis of NMR line shapes and chemical shifts. We discuss alternative interpretations of this observation in terms of FKBP12 function.</p>},
  author       = {Weininger, Ulrich and Modig, Kristofer and Geitner, Anne Juliane and Schmidpeter, Philipp A.M. and Koch, Johanna R. and Akke, Mikael},
  issn         = {0006-2960},
  language     = {eng},
  month        = {01},
  number       = {1},
  pages        = {334--343},
  publisher    = {The American Chemical Society},
  series       = {Biochemistry},
  title        = {Dynamics of Aromatic Side Chains in the Active Site of FKBP12},
  url          = {http://dx.doi.org/10.1021/acs.biochem.6b01157},
  volume       = {56},
  year         = {2017},
}