Heat of reaction between trypsin and soybean trypsin inhibitor

Dobry, Alan; Sturtevant, Julian M.

doi:10.1016/0003-9861(52)90188-4

Cited by 19 publications

(6 citation statements)

References 8 publications

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“…44 Specifically, even if a pK a 8.3 is assumed, at pH 7.5 one estimates at least 50% preservation of the buffer capacity. As the temperature trend for pK a is similar to that of pK (2) (both displaying DH o % 47 6 3 kJ/ mol 43,44 ), it follows that the buffer capacity of 50 mM Tris buffer around pH 7.5 is nearly unaffected by temperature, and, hence, there is very little, if any, contribution from this source to the respective activation enthalpy for the Bz-Gly-Phe hydrolysis (vide infra). All the working solutions were prepared daily before the experimental series were started.…”

Section: Chemicals and Solutionsmentioning

confidence: 81%

“…The working concentration of CPA in routine experiments was 1 nM, and of the substrates 1 mM. For the case of peptide substrate, concentration of the ''active'' (dissociable) product, Phe (with pK (2) $ 9.1/9.3) 43 was in the range of 0-0.5 mM for kinetically evaluated progress curve regions. Thus, the buffer concentration always exceeded the concentration of dissociable Phe species at least 100 times.…”

Section: Chemicals and Solutionsmentioning

confidence: 99%

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Diverse role of conformational dynamics in carboxypeptidase A‐driven peptide and ester hydrolyses: Disclosing the “Perfect Induced Fit” and “Protein Local Unfolding” pathways by altering protein stability

et al. 2011

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Catalytic mechanisms of carboxypeptidase A (CPA) are well known for their diversity and the relative inaccessibility for a decisive comprehension. Recent encouraging attempts through modern computational techniques promoted new challenges for the complementary experimental endeavors. In this work, we have applied the stopped-flow technique and the method of reaction progress curve fitting to extract kinetic parameters for the CPA-catalyzed hydrolyses of smaller (typical) peptide and ester substrates, known for their strong activating/inhibiting impact, thus to which the traditional method of "initial rates" is not applicable. Our approach that innately implies the overall constancy of the affecter (substrate plus "active" product) concentration, made it possible to rigorously determine the physically meaningful "effective" values for the catalytic and Michaelis constants under diverse experimental conditions including variable temperature and urea or trimethylamine N-oxide concentrations. Analysis of the obtained results allowed for: (i) the further substantiation of diverse mechanistic patterns for archetypal specific peptide and ester substrates, (ii) testing and disclosure of intrinsic links between the stabilizing/destabilizing and activating/inhibiting effects for the important model enzyme, CPA, and (iii) tentative explanation of a distinct activating/inhibiting impact of these substrates through the strong specific interaction of their benzyl (Bz) moiety with the substrate binding S(3) subsite of CPA. We have demonstrated that stabilization of CPA either through the interaction with an extra Bz moiety (belonging to another substrate or to the product) leads to the increase of its catalytic power with respect to the specific peptide substrate and to its decrease with respect to the counterpart ester substrate. We conjecture that the catalytic mechanisms operating in these two cases include: (a) the "promoted water" mechanism for the peptide substrate that, seemingly, provides the almost "perfect induced fit" (low-barrier conformational adaptation), and (b) presumably, the "anhydride intermediate" mechanism for the ester substrate that, anyway, requires substantial conformational rearrangement (in fact, "partial or local unfolding") of the protein environment in the course of the rate-determining step.

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Section: Chemicals and Solutionsmentioning

confidence: 81%

Section: Chemicals and Solutionsmentioning

confidence: 99%

Diverse role of conformational dynamics in carboxypeptidase A‐driven peptide and ester hydrolyses: Disclosing the “Perfect Induced Fit” and “Protein Local Unfolding” pathways by altering protein stability

et al. 2011

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“…Consider in greater detail the two reactions in which the amide bonds of asparagine and glutamine are hydrolyzed [65][66][67][68]71]: "0 NHj I = -02C-CH-CH2-CHz-COz +NHt (9) (L-glutamate) LlH r = -21.8 kJ mol-i ; LlG r = -11.7 kJ mol-i. Glycine-lysine -8.8±O.3 [67] As expected, the structurally similar compounds aspargine and glutamine have similar thermodynamic parameters for the hydrolysis reactions.…”

Section: Thermodynamics Of Enzymatic Reactions Catalyzed By Hydrolasesmentioning

confidence: 99%

“…Table 6 shows that the enthalpies of peptide bond hydrolysis range from -5.2 kJ mol-i to -10.7 kJ mol-i ; the mean value is -7.5 kJ mol-i. Analogous determination of the mean enthalpy values for hydrolysis of some other bonds in aqueous solution: ~ -25 kJ mol-i for amide bonds [65][66][67][68]; ~ + 2 kJ mol-i for ester bonds [59-61, 72] and ~ -4 kJ mol-i for thioester bonds [59][60][61].…”

Section: Thermodynamics Of Enzymatic Reactions Catalyzed By Hydrolasesmentioning

confidence: 99%

Thermodynamics of Enzymatic Reactions

Rekharsky¹,

Galchenko²,

Егоров³

et al. 1986

Thermodynamic Data for Biochemistry and Biotechnology

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“…Circuits that today would be considered to be extremely crude and bulky were designed to square, integrate, and record the feedback current. This instrument was used over a period of many years to measure the enthalpy changes in a wide variety of biochemically important processes, including the isothermal denaturation of pepsin by changes in pH; the hydrolysis of peptide bonds (Dobry et al, 1952); the reaction between trypsin and soybean trypsin inhibitor; the hydrolysis of inorganic and organic phosphates; the polymerization and clotting of fibrin monomers (Laskowski et al, 1955); the denaturation of DNA; the reaction between polyriboadenylic acid and polyribouridylic acid; and many others.…”

mentioning

confidence: 99%