Probing backbone hydrogen bonding in PDZ/ligand interactions by protein amide-to-ester mutations

TY - JOUR

T1 - Probing backbone hydrogen bonding in PDZ/ligand interactions by protein amide-to-ester mutations

AU - Pedersen, Søren W

AU - Pedersen, Stine B

AU - Anker, Louise

AU - Hultqvist, Greta

AU - Kristensen, Anders S

AU - Jemth, Per

AU - Strømgaard, Kristian

PY - 2014

Y1 - 2014

N2 - PDZ domains are scaffolding modules in protein-protein interactions that mediate numerous physiological functions by interacting canonically with the C-terminus or non-canonically with an internal motif of protein ligands. A conserved carboxylate-binding site in the PDZ domain facilitates binding via backbone hydrogen bonds; however, little is known about the role of these hydrogen bonds due to experimental challenges with backbone mutations. Here we address this interaction by generating semisynthetic PDZ domains containing backbone amide-to-ester mutations and evaluating the importance of individual hydrogen bonds for ligand binding. We observe substantial and differential effects upon amide-to-ester mutation in PDZ2 of postsynaptic density protein 95 and other PDZ domains, suggesting that hydrogen bonding at the carboxylate-binding site contributes to both affinity and selectivity. In particular, the hydrogen-bonding pattern is surprisingly different between the non-canonical and canonical interaction. Our data provide a detailed understanding of the role of hydrogen bonds in protein-protein interactions.

AB - PDZ domains are scaffolding modules in protein-protein interactions that mediate numerous physiological functions by interacting canonically with the C-terminus or non-canonically with an internal motif of protein ligands. A conserved carboxylate-binding site in the PDZ domain facilitates binding via backbone hydrogen bonds; however, little is known about the role of these hydrogen bonds due to experimental challenges with backbone mutations. Here we address this interaction by generating semisynthetic PDZ domains containing backbone amide-to-ester mutations and evaluating the importance of individual hydrogen bonds for ligand binding. We observe substantial and differential effects upon amide-to-ester mutation in PDZ2 of postsynaptic density protein 95 and other PDZ domains, suggesting that hydrogen bonding at the carboxylate-binding site contributes to both affinity and selectivity. In particular, the hydrogen-bonding pattern is surprisingly different between the non-canonical and canonical interaction. Our data provide a detailed understanding of the role of hydrogen bonds in protein-protein interactions.

U2 - 10.1038/ncomms4215

DO - 10.1038/ncomms4215

M3 - Journal article

C2 - 24477114

SN - 2041-1723

VL - 5

SP - 1

EP - 11

JO - Nature Communications

JF - Nature Communications

M1 - 3215

ER -