Fig. 7.

Functional importance and conservation of solvent gating in DHFR. (A) The rate of hydride transfer, $k_{\mathit{hyd}}$, for selected mutants of ecDHFR. (B) The structure of the N23PP/S148A mutant of ecDHFR (PDB: 3QL0) shows well-supported density for an ordered water in the $2m{F}_o-D{F}_c$ map (blue mesh; 1.5$\sigma$). (C) Structures of human DHFR (PDB: 4M6K and PDB: 2W3M, molecule B) have unmodeled density consistent with partial-occupancy water within 3.5 Å of the N5 nitrogen of FOL and evidence of an alternate rotamer for Phe31 ($m{F}_o-D{F}_c$; green/red mesh; $\pm 3.5\sigma$). A single rotamer is supported for Leu22 in the $2m{F}_o-D{F}_c$ maps (blue mesh; 1.0$\sigma$) suggesting that Phe31 instead serves as the solvent-gating residue in the human enzyme. Although only molecule B is presented for the 2W3M deposited structure, similar features are observed in both protein molecules of the asymmetric unit.