The Energetics of Hydrogen Bonds in Model Systems: Implications for Enzymatic Catalysis

Shan, Shu‐ou; Loh, Stewart N.; Herschlag, Daniel

doi:10.1126/science.272.5258.97

Cited by 268 publications

(232 citation statements)

References 60 publications

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“…The increased stabilization is most likely to be the result of the combined contributions from two known effects on hydrogen bonds exposed to the bulk solvent: (i) Becker et al (61) have elegantly demonstrated that the interior of both DNA and RNA duplexes are devoid of water by showing a significant decrease in acridinium ester hydrolysis inside a complementary double helix compared to mismatched duplexes. Further, Shan et al have shown that the strength of hydrogen bonding is proportional to the dielectric of the solvent (62,63). Enzymatic catalysis is proposed to be achieved as the interior of the enzyme has a much lower dielectric than the surrounding solvent, making the hydrogen bonds stabilizing the transition state stronger inside the active site than free in solution, thereby catalyzing the reaction.…”

Section: Resultsmentioning

confidence: 99%

A Uniform Mechanism Correlating Dangling-End Stabilization and Stacking Geometry

Isaksson

Chattopadhyaya

2005

Biochemistry

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The geometry of the dangling base in 105 published structures (from X-ray/NMR) containing single-stranded overhangs has been analyzed and correlated to the thermodynamic stabilization found (UV) for the corresponding dangling base/closing basepair combination in short oligonucleotides. The study considers most combinations of closing basepairs, sequence and dangling base residue type, attached in both the 3′-and 5′-ends of both DNA and RNA. Linear regression analysis showed a straightforward correlation (R ) 0.873) between the degree of screening for the hydrogen bonds of the closing basepair provided by the dangling base and the resulting thermodynamic stabilization in both DNA and RNA series with dangling ends either at the 3′-or at the 5′-terminus. Regression analysis of only the datasets from RNA gives an improved correlation, R ) 0.934, showing that dangling ends on RNA are more ordered than the dangling ends on DNA, R ) 0.376. This study highlights the gain in the free energy of stabilization owing to the favorable stacking between the dangling nucleobase and the neighboring basepair and the resulting strengthening of the hydrogen bond of the closing basepair. By acting as a hydrophobic cap on the terminal of the DNA or RNA duplex, the dangling-end residue restricts the bulk water access to the terminal basepair, thereby providing it with a microenvironment devoid of water, which consequently enhances its thermodynamic stability, making it energetically comparable to the corresponding internal basepair. Thus, one single structural model consisting of the interplay of the above electrostatic interactions can be used to explain the molecular basis of the observed thermodynamic effects for dangling-end attachment to the 3′-and 5′-ends of both DNA and RNA duplexes, which is a key step toward accurate dangling-end effect prediction.Unpaired terminal nucleotides naturally occur in various biologically important RNA structures: The acceptor stem in tRNA (1-3) has a characteristic 3′-NCCA nucleobase overhang that determines its structure, stability, and function. The dangling ends neighboring the codon-anticodon pair have also been found to stabilize the interaction between tRNA and mRNA (4, 5) for protein synthesis in the ribosomal machinery. The efficiency of down regulation of gene expression by RNA interference (RNAi) has been shown to be dependent on a two to three nucleotide overhang (6-9). Similarly, the structures of pseudoknots (10) are stabilized by a complicated interplay between single-stranded and double-stranded motifs. A multitude of literature on the stabilizing interactions of native single-stranded oligonucleotides is available (9,(11)(12)(13)(14).Accurate prediction of nucleic acid secondary structure and stability from its primary sequence is a key interest for probe binding optimization in biotechnological applications such as microarrays (15) and molecular beacons (16). To develop an accurate algorithm (14,(17)(18)(19)(20)(21)(22)(23)(24) for predicting the thermodynamics of hybridi...

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Section: Resultsmentioning

confidence: 99%

A Uniform Mechanism Correlating Dangling-End Stabilization and Stacking Geometry

Isaksson

Chattopadhyaya

2005

Biochemistry

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“…Extensive studies and debates have appeared in the literature since the idea of low barrier hydrogen bond (LBHB or SSHB) was first proposed (12)(13)(14)(15). Two of the key questions are the strength of the LBHB and its contribution to the rate enhancement of the reaction catalyzed by the enzyme.…”

Section: Resultsmentioning

confidence: 99%

The Catalytic Role of Aspartate in a Short Strong Hydrogen Bond of the Asp274–His32 Catalytic Dyad in Phosphatidylinositol-specific Phospholipase C Can Be Substituted by a Chloride Ion

Zhao¹,

Liao

Tsai

2004

Journal of Biological Chemistry

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Phosphatidylinositol-specific phospholipase C from Bacillus thuringiensis catalyzes the cleavage of the phosphorus-oxygen bond in phosphatidylinositol. The focus of this work is to dissect the roles of the carboxylate side chain of Asp 274 in the Asp 274 -His 32 dyad, where a short strong hydrogen bond (SSHB) was shown to exist based on NMR criteria. A regular hydrogen bond (HB) was observed in D274N, and no low field proton resonance was detected for D274E and D274A. Comparison of the activity of wild type, D274N, and D274A suggested that the regular HB contributes significantly (ϳ4 kcal/ mol) to catalysis, whereas the SSHB contributes only an additional 2 kcal/mol. The mutant D274E displays high activity similar to wild type, suggesting that the negative charge is sufficient for the catalytic role of Asp 274 . To further support this interpretation and rule out possible contribution of regular HB or SSHB in D274E, we showed that the activity of D274G can be rescued by exogenous chloride ions to a level comparable with that of D274E. Comparison between different anions suggested that the ability of an anion to rescue the activity is due to the size and the charge of the anion not the property as a HB acceptor. In conclusion, a major fraction of the functional role of Asp 274 in the Asp 274 -His 32 dyad can be attributed to a negative charge (as in D274E and D274G-Cl ؊ ), and the SSHB in the wild type enzyme provides minimal contribution to catalysis. These results represent novel insight for an Asp-His catalytic dyad and for the mechanism of phosphatidylinositolspecific phospholipase C.

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“…A second one is the increase of hydrogen bond strength induced by decreasing the dielectric constant and concomitant formation of transient charges (16). It appears that this aspect of MCADH is an example for what was suggested recently by Shan and Herschlag (13,55). A third concept is the combined effect of H-bonds, dielectric changes, and charge rearrangements on the redox potentials of substrate and flavin (i.e., the redox partners, 43).…”

Section: Discussionmentioning

confidence: 99%

“…A crucial issue in the dehydrogenation reaction of acylCoA dehydrogenase is the mode and extent of activation of RC-H. For efficient catalysis, current concepts assume that the microscopic pK a values of two partners involved in the transfer of one H + between them have similar values (12,13). The microscopic pK a of Glu376sCOO -recently has been determined to be ≈6 in uncomplexed enzyme (14).…”

mentioning

confidence: 99%

Mechanism of Activation of Acyl-CoA Substrates by Medium Chain Acyl-CoA Dehydrogenase: Interaction of the Thioester Carbonyl with the Flavin Adenine Dinucleotide Ribityl Side Chain

et al. 1998

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The flavin adenine dinucleotide (FAD) cofactor of pig kidney medium-chain specific acylcoenzyme A (CoA) dehydrogenase (MCADH) has been replaced by ribityl-3′-deoxy-FAD and ribityl-2′-deoxy-FAD. 3′-Deoxy-FAD-MCADH has properties very similar to those of native MCADH, indicating that the FAD-ribityl side-chain 3′-OH group does not play any particular role in cofactor binding or catalysis. 2′-Deoxy-FAD-MCADH was characterized using the natural substrate C 8 CoA as well as various substrate and transition-state analogues. Substrate dehydrogenation in 2′-deoxy-FAD-MCADH is ≈1.5 × 10 7 -fold slower than that of native MCADH, indicating that disruption of the hydrogen bond between 2′-OH and substrate thioester carbonyl leads to a substantial transition-state destabilization equivalent to ≈38 kJ mol -1 . The RC-H microscopic pK a of the substrate analogue 3S-C 8 CoA, which undergoes R-deprotonation on binding to MCADH, is lowered from ≈16 in the free state to ≈11 ((0.5) when bound to 2′-deoxy-FAD-MCADH. This compares with a decrease of the same pK a to ≈5 in the complex with unmodified hwtMCADH, which corresponds to a pK shift of ≈11 pK units, i.e., ≈65 kJ mol -1 [Vock, P., Engst, S., Eder, M., and Ghisla, S. (1998) Biochemistry 37, 1848-1860]. The difference of this effect of ≈6 pK units (≈35 kJ mol -1 ) between MCADH and 2′-deoxy-FAD-MCADH is taken as the level of stabilization of the substrate carbanionic species caused by the interaction with the FAD-2′-OH. This energetic parameter derived from the kinetic experiments (stabilization of transition state) is in agreement with those obtained from static experiments (lowering of RC-H microscopic pK a of analogue, i.e., stabilization of anionic transition-state analogue). The contributions of the two single H-bonds involved in substrate activation (Glu376amide-N-H and ribityl-2′-OH) thus appear to behave additively toward the total effect. The crystal structures of native pMCADH and of 2′-deoxy-FAD-MCADH complexed with octanoyl-CoA/octenoyl-CoA show unambiguously that the FAD cofactor and the substrate/product bind in an identical fashion, implying that the observed effects are mainly due to (the absence of) the FAD-ribityl-2′-OH hydrogen bond. The large energy associated with the 2′-OH hydrogen bond interaction is interpreted as resulting from the changes in charge and the increased hydrophobicity induced by binding of lipophilic substrate. This is the first example demonstrating the direct involvement of a flavin cofactor side chain in catalysis.

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The Energetics of Hydrogen Bonds in Model Systems: Implications for Enzymatic Catalysis

Cited by 268 publications

References 60 publications

A Uniform Mechanism Correlating Dangling-End Stabilization and Stacking Geometry

A Uniform Mechanism Correlating Dangling-End Stabilization and Stacking Geometry

The Catalytic Role of Aspartate in a Short Strong Hydrogen Bond of the Asp274–His32 Catalytic Dyad in Phosphatidylinositol-specific Phospholipase C Can Be Substituted by a Chloride Ion

Mechanism of Activation of Acyl-CoA Substrates by Medium Chain Acyl-CoA Dehydrogenase: Interaction of the Thioester Carbonyl with the Flavin Adenine Dinucleotide Ribityl Side Chain

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