Probing
Water Environment of Trp59 in Ribonuclease
T1: Insight of the Structure–Water Network Relationship
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Abstract
In
this study, we used the tryptophan analogue, (2,7-aza)Trp, which
exhibits water catalyzed proton transfer isomerization among N(1)-H,
N(7)-H, and N(2)-H isomers, to probe the water environment of tryptophan-59
(Trp59) near the connecting loop region of ribonuclease Tl (RNase
T1) by replacing the tryptophan with (2,7-aza)Trp. The resulting (2,7-aza)Trp59
triple emission bands and their associated relaxation dynamics, together
with relevant data of 7-azatryptophan and molecular dynamics (MD)
simulation, lead us to propose two Trp59 containing conformers in
RNase T1, namely, the loop-close and loop-open forms. Water is rich
in the loop-open form around the proximity of (2,7-aza)Trp59, which
catalyzes (2,7-aza)Trp59 proton transfer in the excited state, giving
both N(1)-H and N(7)-H isomer emissions. The existence of N(2)-H isomer
in the loop-open form, supported by the MD simulation, is mainly due
to the specific hydrogen bonding between N(2)-H proton and water molecule
that bridges N(2)-H and the amide oxygen of Pro60, forming a strong
network. The loop-close form is relatively tight in space, which squeezes
water molecules out of the interface of α-helix and β2
strand, joined by the connecting loop region; accordingly, the water-scant
environment leads to the sole existence of the N(1)-H isomer emission.
MD simulation also points out that the Trp-water pairs appear to preferentially
participate in a hydrogen bond network incorporating polar amino acid
moieties on the protein surface and bulk waters, providing the structural
dynamic features of the connecting loop region in RNase T1