Phosphonamidates as transition-state analog inhibitors of thermolysin

Bartlett, Paul A.; Marlowe, Charles K.

doi:10.1021/bi00289a002

Cited by 287 publications

(223 citation statements)

References 79 publications

Supporting

Mentioning

212

Contrasting

Unclassified

Order By: Relevance

“…9 Peptide analogues containing a phosphonate linkage, when compared with substrates for the zinc protease thermolysin, show a slope of 1.05. 7 A peptide analogue with a phosphonamidate linkage yield a slope of 1.03 when alterations were made at a catalytic arginine in carboxypeptidase A. 13 The linear correlations of U>v inhibition and catalytic changes, resulting from alterations made to RNase A at position 41, show slopes that are much more shallow, between 0.25 and 0.36 ( Figure 2).…”

Section: Is Uridine 2′3′-cyclic Vanadate a Transition State Analogue?mentioning

confidence: 99%

“…7,10,12,13 The failure of pentavalent organo-vanadates as transition state analogues for phosphoryl transfer reactions is explicable. The location and charge of the vanadyl oxygens differ significantly from those of the corresponding phosphoryl oxygens in the enzymic transition state.…”

Section: Organo-vanadates As Mimics Of Transition Statesmentioning

confidence: 99%

“…Bartlett has therefore argued that a more appropriate criteria for assessing transition state analogy is that the relative affinity of an enzyme for the chemical transition state (given by k cat /K m ) and for a putative transition state analogue (given by 1/K i ) be constant over a range of values. 7,8 In other words, Δlog(K m /k cat ) = ΔlogK i .…”

Section: Introductionmentioning

confidence: 99%

“…This first method has been applied extensively to proteases, using an aldehyde, phosphonate, or phosphonamidate group in place of the scissile amide in a peptidyl analogue. 7,[9][10][11][12] The other method involves systematic alteration of the active site. This second method is more generally applicable, as it does not require the enzyme to accommodate a variety of substrates.…”

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Pentavalent Organo-Vanadates as Transition State Analogues for Phosphoryl Transfer Reactions

Messmore

Raines

2000

J. Am. Chem. Soc.

View full text Add to dashboard Cite

Pentavalent organo-vanadates have been put forth as transition state analogues for a variety of phosphoryl transfer reactions. In particular, uridine 2′,3′-cyclic vanadate (U>v) has been proposed to resemble the transition state during catalysis by ribonuclease A (RNase A). Here, this hypothesis is tested. Lys41 of RNase A is known to donate a hydrogen bond to a nonbridging phosphoryl oxygen in the transition state during catalysis. Site-directed mutagenesis and semisynthesis were used to create enzymes with natural and nonnatural amino acid residues at position 41. These variants differ by 10 5 -fold in their k cat /K m values for catalysis, but <40-fold in their K i values for inhibition of catalysis by U>v. Plots of logK i vs log(K m /k cat ) for three distinct substrates [poly(cytidylic acid), uridine 3′-(p-nitrophenyl phosphate), and cytidine 2′,3′-cyclic phosphate] have slopes that range from 0.25 and 0.36. These plots would have a slope of unity if U>v were a perfect transition state analogue. Values of K i for U>v correlate weakly with the equilibrium dissociation constant for the enzymic complexes with substrate or product, indicating that U>v bears some resemblance to the substrate and product as well as the transition state. Thus, U>v is a transition state analogue for RNase A, but only a marginal one. This finding indicates that a pentavalent organo-vanadate cannot necessarily be the basis for a rigorous analysis of the transition state for a phosphoryl transfer reaction.

show abstract

Section: Is Uridine 2′3′-cyclic Vanadate a Transition State Analogue?mentioning

confidence: 99%

Section: Organo-vanadates As Mimics Of Transition Statesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Pentavalent Organo-Vanadates as Transition State Analogues for Phosphoryl Transfer Reactions

Messmore

Raines

2000

J. Am. Chem. Soc.

View full text Add to dashboard Cite

show abstract

“…Substrate analogs containing phosphonate, phosphonoamidate, and phosphinate groups have been used as effective noncovalent inhibitors of aspartylproteases (24) and metalloproteases (25), whereas phosphonate and phosphinate bisubstrate inhibitors have been used to inhibit farnesyltransferase (26). Of particular interest to the enzymatic mechanism of FAH are studies indicating that the tetrahedral geometry associated with the phosphorus groups in these types of inhibitors closely approximates both geometric and electronic aspects of the transition state (24,27,28). Thus, the effectiveness of these compounds as inhibitors is presumably associated with similarities to the high energy reaction intermediates stabilized during catalysis.…”

mentioning

confidence: 99%

Mechanistic Inferences from the Crystal Structure of Fumarylacetoacetate Hydrolase with a Bound Phosphorus-based Inhibitor

Bateman

Bhanumoorthy

Witte

et al. 2001

Journal of Biological Chemistry

View full text Add to dashboard Cite

Thermolysin-inhibitor binding: Effect of the His231 → Ala mutation on the relative affinities of thiolate versus phosphoramidate inhibitors—a model theoretical investigation incorporating acontinuum reaction field hydration model

Gresh

Roques

1997

Biopolymers

View full text Add to dashboard Cite

A series of site‐directed mutagenesis experiments have been performed in our laboratory, bearing on the Zn2+ metalloprotease from Bacillus stearothermophillus, a protein very closely related to thermolysin (TLN), and their consequences on the binding of inhibitors belonging to the thiolate, carboxylate, hydroxamate, and phosphoramidate classes of compounds [A. Beaumont et al. (1995), J. Biol. Chem., Vol. 270, pp. 16803–16808]. An unexpected result related to the mutation of protonated His231, which resides at the entrance of the enzymatic cavity, into an Ala residue, shown to leave the binding affinities of thiolate inhibitors unaffected, but to abolish almost completely those of the phosphoramidate inhibitors. In order to account for such contrasting differential responses, we have undertaken a theoretical study of the comparative binding affinities of a representative thiol inhibitor, thiorphan (I) and a phosphoramidate (II), to a model of the enzymatic cavity of both the native and the mutated protein, encompassing up to 27 residues. The computations of inter‐ (ΔE) and intramolecular interactions were done with the sum of interactions between fragments ab initio computed procedure [N. Gresh et al. (1984), Theoret. Chim. Acta. Vol. 66, pp. 1–20; (1985), Theoret. Chim. Acta, Vol. 67, pp. 11–32; (1986), Int. J. Quant. Chem., Vol. 29, pp. 101–118; (1994), in Modelling the Hydrogen Bond, American Chemical Society Symposia, Vol. 569, D. A. Smith, Ed., American Chemical Society, pp. 82–112; N. Gresh (1995), J. Comput. Chem., Vol. 16, 856–882]. The energy balances for the His231 → Ala mutation incorporate a solvation energy contribution, computed using the Langlet et al. Continuum procedure [J. Langlet et al. (1988), J. Phys. Chem., Vol. 92, pp. 1617–1631], and this term was shown to play a decisive role in the comparative energy balances of the thiolate vs phosphoramidate inhibitors. Whereas the resolvation energy was able to compensate for the loss of ΔE induced by the H231 A mutation for the complex of I, thus accounting for its insensitivity to the mutation, such a compensation did not occur in the case of II, leading in its complexes to an overall loss of about 7 kcal/mole in the mutated cavity with respect to the native one. These results emphasized the critical role played by solvation in enzyme‐inhibitor recognition processses. © 1997 John Wiley & Sons, Inc.

show abstract

Phosphonamidates as transition-state analog inhibitors of thermolysin

Cited by 287 publications

References 79 publications

Pentavalent Organo-Vanadates as Transition State Analogues for Phosphoryl Transfer Reactions

Pentavalent Organo-Vanadates as Transition State Analogues for Phosphoryl Transfer Reactions

Mechanistic Inferences from the Crystal Structure of Fumarylacetoacetate Hydrolase with a Bound Phosphorus-based Inhibitor

Thermolysin-inhibitor binding: Effect of the His231 → Ala mutation on the relative affinities of thiolate versus phosphoramidate inhibitors—a model theoretical investigation incorporating acontinuum reaction field hydration model

Contact Info

Product

Resources

About