TY - JOUR
T1 - Phosphorylation releases constraints to domain motion in ERK2
AU - Xiao, Yao
AU - Lee, Thomas
AU - Latham, Michael Parker
AU - Warner, Lisa Rose
AU - Tanimoto, Akiko
AU - Pardi, Arthur
AU - Ahn, Natalie G.
PY - 2014/2/18
Y1 - 2014/2/18
N2 - Protein motions control enzyme catalysis through mechanisms that are incompletely understood. Here NMR 13C relaxation dispersion experiments were used to monitor changes in side-chain motions that occur in response to activation by phosphorylation of the MAP kinase ERK2. NMR data for the methyl side chains on Ile, Leu, and Val residues showed changes in conformational exchange dynamics in the microsecond-to-millisecond time regime between the different activity states of ERK2. In inactive, unphosphorylated ERK2, localized conformational exchange was observed among methyl side chains, with little evidence for coupling between residues. Upon dual phosphorylation by MAP kinase kinase 1, the dynamics of assigned methyls in ERK2 were altered throughout the conserved kinase core, including many residues in the catalytic pocket. The majority of residues in active ERK2 fit to a single conformational exchange process, with kex ̃ 300 s-1 (kAB ̃ 240 s-1/kBA ̃ 60 s-1) and pA/pB ̃ 20%/80%, suggesting global domain motions involving interconversion between two states. A mutant of ERK2, engineered to enhance conformational mobility at the hinge region linking the N- and C-terminal domains, also induced two-state conformational exchange throughout the kinase core, with exchange properties of kex ̃ 500 s-1 (kAB ̃ 15 s-1/kBA ̃ 485 s-1) and pA/pB ̃ 97%/3%. Thus, phosphorylation and activation of ERK2 lead to a dramatic shift in conformational exchange dynamics, likely through release of constraints at the hinge.
AB - Protein motions control enzyme catalysis through mechanisms that are incompletely understood. Here NMR 13C relaxation dispersion experiments were used to monitor changes in side-chain motions that occur in response to activation by phosphorylation of the MAP kinase ERK2. NMR data for the methyl side chains on Ile, Leu, and Val residues showed changes in conformational exchange dynamics in the microsecond-to-millisecond time regime between the different activity states of ERK2. In inactive, unphosphorylated ERK2, localized conformational exchange was observed among methyl side chains, with little evidence for coupling between residues. Upon dual phosphorylation by MAP kinase kinase 1, the dynamics of assigned methyls in ERK2 were altered throughout the conserved kinase core, including many residues in the catalytic pocket. The majority of residues in active ERK2 fit to a single conformational exchange process, with kex ̃ 300 s-1 (kAB ̃ 240 s-1/kBA ̃ 60 s-1) and pA/pB ̃ 20%/80%, suggesting global domain motions involving interconversion between two states. A mutant of ERK2, engineered to enhance conformational mobility at the hinge region linking the N- and C-terminal domains, also induced two-state conformational exchange throughout the kinase core, with exchange properties of kex ̃ 500 s-1 (kAB ̃ 15 s-1/kBA ̃ 485 s-1) and pA/pB ̃ 97%/3%. Thus, phosphorylation and activation of ERK2 lead to a dramatic shift in conformational exchange dynamics, likely through release of constraints at the hinge.
UR - http://www.scopus.com/inward/record.url?scp=84894379546&partnerID=8YFLogxK
U2 - 10.1073/pnas.1318899111
DO - 10.1073/pnas.1318899111
M3 - Article
C2 - 24550275
AN - SCOPUS:84894379546
SN - 0027-8424
VL - 111
SP - 2506
EP - 2511
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 7
ER -