Double Mutant of Chymotrypsin Inhibitor 2 Stabilized through Increased Conformational Entropy
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Double Mutant of Chymotrypsin Inhibitor 2 Stabilized through Increased Conformational Entropy. / Gavrilov, Yulian; Kümmerer, Felix; Orioli, Simone; Prestel, Andreas; Lindorff-Larsen, Kresten; Teilum, Kaare.
I: Biochemistry, Bind 61, Nr. 3, 2022, s. 160-170.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - Double Mutant of Chymotrypsin Inhibitor 2 Stabilized through Increased Conformational Entropy
AU - Gavrilov, Yulian
AU - Kümmerer, Felix
AU - Orioli, Simone
AU - Prestel, Andreas
AU - Lindorff-Larsen, Kresten
AU - Teilum, Kaare
PY - 2022
Y1 - 2022
N2 - The conformational heterogeneity of a folded protein can affect not only its function but also stability and folding. We recently discovered and characterized a stabilized double mutant (L49I/I57V) of the protein CI2 and showed that state-of-the-art prediction methods could not predict the increased stability relative to the wild-type protein. Here, we have examined whether changed native-state dynamics, and resulting entropy changes, can explain the stability changes in the double mutant protein, as well as the two single mutant forms. We have combined NMR relaxation measurements of the ps-ns dynamics of amide groups in the backbone and the methyl groups in the side chains with molecular dynamics simulations to quantify the native-state dynamics. The NMR experiments reveal that the mutations have different effects on the conformational flexibility of CI2: a reduction in conformational dynamics (and entropy estimated from this) of the native state of the L49I variant correlates with its decreased stability, while increased dynamics of the I57V and L49I/I57V variants correlates with their increased stability. These findings suggest that explicitly accounting for changes in native-state entropy might be needed to improve the predictions of the effect of mutations on protein stability.
AB - The conformational heterogeneity of a folded protein can affect not only its function but also stability and folding. We recently discovered and characterized a stabilized double mutant (L49I/I57V) of the protein CI2 and showed that state-of-the-art prediction methods could not predict the increased stability relative to the wild-type protein. Here, we have examined whether changed native-state dynamics, and resulting entropy changes, can explain the stability changes in the double mutant protein, as well as the two single mutant forms. We have combined NMR relaxation measurements of the ps-ns dynamics of amide groups in the backbone and the methyl groups in the side chains with molecular dynamics simulations to quantify the native-state dynamics. The NMR experiments reveal that the mutations have different effects on the conformational flexibility of CI2: a reduction in conformational dynamics (and entropy estimated from this) of the native state of the L49I variant correlates with its decreased stability, while increased dynamics of the I57V and L49I/I57V variants correlates with their increased stability. These findings suggest that explicitly accounting for changes in native-state entropy might be needed to improve the predictions of the effect of mutations on protein stability.
KW - MAGNETIC-RESONANCE RELAXATION
KW - SIDE-CHAIN DYNAMICS
KW - MODEL-FREE APPROACH
KW - HYDROPHOBIC CORE MUTATIONS
KW - DEUTERIUM SPIN PROBES
KW - TRANSITION-STATE
KW - MOLECULAR SIMULATIONS
KW - HYDROGEN-EXCHANGE
KW - ORDER PARAMETERS
KW - PROTEINS
U2 - 10.1021/acs.biochem.1c00749
DO - 10.1021/acs.biochem.1c00749
M3 - Journal article
C2 - 35019273
VL - 61
SP - 160
EP - 170
JO - Biochemistry
JF - Biochemistry
SN - 0006-2960
IS - 3
ER -
ID: 291215544