Short, strong hydrogen bonds on enzymes: NMR and mechanistic studies

The lengths of short, strong hydrogen bonds (SSHBs) on enzymes have been determined with high precision (+/-0.05 A) from the chemical shifts (delta), and independently from the D/H fractionation factors (phi) of the highly deshielded protons involved. These H-bond lengths agree well with each other and with those found by protein X-ray crystallography, within the larger errors' of the latter method (+/-0.2 to +/-0.8 Angstrom) [Proteins 35 (1999) 275]. A model dihydroxy-naphthalene compound shows a SSHB of 2.54 +/- 0.04 Angstrom based on delta = 17.7 ppm and phi = 0.56 0.04, in agreement with the high resolution X-ray distance of 2.55 +/- 0.06 Angstrom. On ketosteroid isomerase, a SSHB is found (2.50 +/- 0.02 Angstrom), based on delta = 18.2 ppm. and phi = 0.34, from Tyr-14 to the 3-O- of estradiol, an analog of the enolate intermediate. Its strength is similar to7 kcal/mol. On triosephosphate isomerase, SSHBs are found from Glu-165 to the 1-NOH of phosphoglycolohydroxamic acid (PGH), an analog of the enolic intermediate (2.55 +/- 0.05 Angstrom), and from His-95 to the enolic-O- of PGH (2.62 +/- 0.02 Angstrom). In the methylglyoxal synthase-PGH complex, a SSHB (2.51 +/- 0.02 Angstrom) forms between Asp-71 and the NOH of PGH with a strength of greater than or equal to4.7 kcal/mol. When serine proteases bind mechanism-based inhibitors which form tetrahedral Ser-adducts analogous to the tetrahedral intermediates in catalysis, the Asp...His H-bond of the catalytic triad becomes a SSHB [Proc. Natl Acad. Sci. USA 95 (1998) 14664], 2.49-2.63 Angstrom in length. Similarly, on the serine-esterase, butyrylcholinegterase complexed with the mechanism-based inhibitor. m-(N,N,N,-trimethylammonio)-2,2,2-trifluoroacetophenone, a SSHB. forms between Glu-327 and His-438 of the catalytic. triad, 2.61 +/- 0.04 Angstrom in length, based on delta = 18.1 ppm and sigma = 0.65 +/- 0.10. Very similar results are obtained with (human) acetylcholinesterase. The strength of this SSHB is at least 4.9 kcal/mol. (C) 2002 Elsevier Science B.V. All rights reserved.