4. Structure for catalysis

Author(s):  
Paul Engel

‘Structure for catalysis’ details the various patterns of enzyme mechanism and the various structural features helping to achieve catalysis. One of the striking features of enzyme catalysis is substrate specificity. In the lock-and-key hypothesis, the enzyme is viewed as a precisely shaped lock and only the right key, the substrate, can fit and turn it. The lock-and-key combination is the enzyme–substrate complex. A crucial ingredient of the enzyme’s equipment for achieving outstanding catalysis is the ‘catalytic groups’.

1966 ◽  
Vol 44 (3) ◽  
pp. 331-337 ◽  
Author(s):  
J. Lyndal York ◽  
G. A. LePage

The kinetic constants Km and Vmax were determined for the deamination by adenosine deaminase of a series of analogues of adenosine containing "fraudulent" sugars. The configuration of the 2′-hydroxyl was found to be important for the binding of enzyme and substrate. The largest effect of changes in sugar structure was on the rate of breakdown of the enzyme–substrate complex to form products, i.e. Vmax. The nature of the configuration in the 3′-position was not important if the 2′-hydroxyl was trans to the glycosidic linkage; however, if the steric arrangement of the 2′-hydroxyl was cis to the glycosidic linkage, then Vmax showed a marked dependence on the nature of the 3′-substituent and its configuration. For instance, Vmax values were for arabinosyl adenine < 3′-deoxyarabinosyl adenine <lyxosyl adenine. 6-N-methyladenosine was found to be a competitive inhibitor of adenosine deaminase, with a Ki of 2 × 10−6M.


2020 ◽  
Author(s):  
Vahe Galstyan ◽  
Kabir Husain ◽  
Fangzhou Xiao ◽  
Arvind Murugan ◽  
Rob Phillips

Key enzymatic processes in biology use the nonequilibrium error correction mechanism called kinetic proofreading to enhance their specificity. Kinetic proofreading typically requires several dedicated structural features in the enzyme, such as a nucleotide hydrolysis site and multiple enzyme–substrate conformations that delay product formation. Such requirements limit the applicability and the adaptability of traditional proofreading schemes. Here, we explore an alternative conceptual mechanism of error correction that achieves delays between substrate binding and subsequent product formation by having these events occur at distinct physical locations. The time taken by the enzyme–substrate complex to diffuse from one location to another is leveraged to discard wrong substrates. This mechanism does not require dedicated structural elements on the enzyme, making it easier to overlook in experiments but also making proofreading tunable on the fly. We discuss how tuning the length scales of enzyme or substrate concentration gradients changes the fidelity, speed and energy dissipation, and quantify the performance limitations imposed by realistic diffusion and reaction rates in the cell. Our work broadens the applicability of kinetic proofreading and sets the stage for the study of spatial gradients as a possible route to specificity.


1997 ◽  
Vol 62 (8) ◽  
pp. 1150-1158 ◽  
Author(s):  
Milan Šolc

The free intramolecular energy flow can be restricted by the presence of a heavy atom in the molecule. As a result of this restriction, adsorbed molecules bonded on the metal surface and/or substrate molecules in the enzyme-substrate complex with a metal atom near the binding site can have a higher vibrational energy than the surroundings. The reaction rate is then enhanced by this energy localization.


2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Serina L Robinson ◽  
Megan D Smith ◽  
Jack E Richman ◽  
Kelly G Aukema ◽  
Lawrence P Wackett

Abstract Enzymes in the thiolase superfamily catalyze carbon–carbon bond formation for the biosynthesis of polyhydroxyalkanoate storage molecules, membrane lipids and bioactive secondary metabolites. Natural and engineered thiolases have applications in synthetic biology for the production of high-value compounds, including personal care products and therapeutics. A fundamental understanding of thiolase substrate specificity is lacking, particularly within the OleA protein family. The ability to predict substrates from sequence would advance (meta)genome mining efforts to identify active thiolases for the production of desired metabolites. To gain a deeper understanding of substrate scope within the OleA family, we measured the activity of 73 diverse bacterial thiolases with a library of 15 p-nitrophenyl ester substrates to build a training set of 1095 unique enzyme–substrate pairs. We then used machine learning to predict thiolase substrate specificity from physicochemical and structural features. The area under the receiver operating characteristic curve was 0.89 for random forest classification of enzyme activity, and our regression model had a test set root mean square error of 0.22 (R2 = 0.75) to quantitatively predict enzyme activity levels. Substrate aromaticity, oxygen content and molecular connectivity were the strongest predictors of enzyme–substrate pairing. Key amino acid residues A173, I284, V287, T292 and I316 in the Xanthomonas campestris OleA crystal structure lining the substrate binding pockets were important for thiolase substrate specificity and are attractive targets for future protein engineering studies. The predictive framework described here is generalizable and demonstrates how machine learning can be used to quantitatively understand and predict enzyme substrate specificity.


2001 ◽  
Vol 56 (11-12) ◽  
pp. 1015-1021 ◽  
Author(s):  
Dámaso Hornero-Méndez ◽  
María Isabel Mínguez-Mosquera

Abstract The in vitro properties of semi-purified chlorophyllase (chlorophyll-chlorophyllido hy­drolase, EC 3.1.1.14) from Capsicum annuum fruits have been studied. The enzym e showed an optimum of activity at pH 8.5 and 50 °C. Substrate specificity was studied for chlorophyll (Chi) a, Chi b, pheophytin (Phe) a and Phe b, with Km values of 10.70, 4.04, 2.67 and 6.37 μᴍ respectively. Substrate inhibition was found for Phe b at concentrations higher than 5 μᴍ. Chlorophyllase action on Chi a' and Chi b' was also studied but no hydrolysis was observed, suggesting that the mechanism of action depends on the configuration at C-132 in the chloro­ phyll molecule, with the enzyme acting only on compounds with R132 stereochemistry. The effect of various metals (Mg2+, Hg2+, Cu2+, Zn2+, Co , Fe2+ and Fe3+) was also investigated, and a general inhibitory effect was found, this being more marked for Hg2+ and Fe2+. Func­tional groups such as -SH and -S-S-seem ed to participate in the formation o f the enzyme-substrate complex. Chelating ion and the carbonyl group at C3 appeared to be important in substrate recognition by the enzyme. The method for measuring Chlase activity, including HPLC separation of substrate and product, has been optimized.


1990 ◽  
Vol 272 (3) ◽  
pp. 613-619 ◽  
Author(s):  
R M Gibson ◽  
H Christensen ◽  
S G Waley

Two single mutants and the corresponding double mutant of beta-lactamase I from Bacillus cereus 569/H were constructed and their kinetics investigated. The mutants have Lys-73 replaced by arginine (K73R), or Glu-166 replaced by aspartic acid (E166D), or both (K73R + E166D). All four rate constants in the acyl-enzyme mechanism were determined for the E166D mutant by the methods described by Christensen, Martin & Waley [(1990) Biochem. J. 266, 853-861]. Both the rate constants for acylation and deacylation for the hydrolysis of benzylpenicillin were decreased about 2000-fold in this mutant. In the K73R mutant, and in the double mutant, the rate constants for acylation were decreased about 100-fold and 10,000-fold respectively. All three mutants also had lowered values for the rate constants for the formation and dissociation of the non-covalent enzyme-substrate complex. The specificities of the mutants did not differ greatly from those of wild-type beta-lactamase, but the hydrolysis of cephalosporin C by the K73R mutant gave ‘burst’ kinetics.


2012 ◽  
Vol 40 (3) ◽  
pp. 515-521 ◽  
Author(s):  
David R. Glowacki ◽  
Jeremy N. Harvey ◽  
Adrian J. Mulholland

One of the most controversial questions in enzymology today is whether protein dynamics are significant in enzyme catalysis. A particular issue in these debates is the unusual temperature-dependence of some kinetic isotope effects for enzyme-catalysed reactions. In the present paper, we review our recent model [Glowacki, Harvey and Mulholland (2012) Nat. Chem. 4, 169–176] that is capable of reproducing intriguing temperature-dependences of enzyme reactions involving significant quantum tunnelling. This model relies on treating multiple conformations of the enzyme–substrate complex. The results show that direct ‘driving’ motions of proteins are not necessary to explain experimental observations, and show that enzyme reactivity can be understood and accounted for in the framework of transition state theory.


2008 ◽  
Vol 9 (3-4) ◽  
pp. 231-243 ◽  
Author(s):  
C. Fenollar-Ferrer ◽  
V. Carnevale ◽  
S. Raugei ◽  
P. Carloni

HIV-1 integrase is the viral enzyme responsible for the insertion of the viral DNA into the host cell chromosome. This process occurs through two distinct biochemical reactions: the 3′-processing of the viral DNA and the transesterification reaction. Because experimental structural information on the reaction intermediate is not available, several molecular models have been developed. Unfortunately, none of the models of the enzyme–substrate complex is fully consistent with the available molecular biological data. We have constructed a new theoretical model based on mutagenesis experiments and cross-linking data, using a relatively accurate computational setup. The structural features of the model along with its limitations are discussed here.


Author(s):  
Gary W. Morrow

It is not essential to have a background in enzymology or biochemistry to gain at least an introductory-level understanding of many biosynthetic processes, so this book does not deal with enzymology or enzyme structure or function in any significant way, even though much of the chemistry we will be examining depends almost entirely on enzyme catalysis. Nevertheless, we will refer to enzyme catalysis and the names of specific enzymes throughout the text as we examine biosynthetic processes and reactions in significant detail. So what exactly are enzymes? Simply put, enzymes are naturally occurring proteins that catalyze various biochemical reactions in living systems. As we will see, many of the reactions they catalyze are familiar organic reactions, but have specific purposes and target structures. Generally speaking, enzymes catalyze organic reactions by lowering transition state energies or raising ground state energies of reactants in much the same way as nonenzymatic catalysts in laboratory chemical reactions, though in the case of enzyme catalysis, rate enhancements of as much as 1023 have been reported, far exceeding rate enhancements currently achievable by conventional chemical means. Understanding the interaction of enzymes and substrates (reactants) to form an enzyme–substrate complex (E–S complex) is fundamental to having some appreciation for how enzymes carry out their work. While overly simplistic, the “lock-and-key” model of enzyme–substrate interaction provides an intuitive context for understanding the remarkable substrate specificity of enzyme-mediated reactions. Thus, so-called enzyme active sites or binding sites (the “lock”) will only accept certain specific substrate structures (the “key”), with shape, conformation, intermolecular forces, and other factors determining the lock-and-key fit. Enzymes not only catalyze specific kinds of reactions, they can act specifically on certain compounds or classes of compounds or functional groups, often showing remarkable selectivity and stereospecificity, especially in the recognition and/or introduction of chirality centers in organic molecules. In terms of nomenclature, enzyme names always end with an ase suffix and are typically named in accordance with the substrate they act upon and/or the kind of reaction process they catalyze.


Acta Naturae ◽  
2014 ◽  
Vol 6 (3) ◽  
pp. 52-65 ◽  
Author(s):  
V. V. Koval ◽  
D. G. Knorre ◽  
O. S. Fedorova

The purpose of the present review is to summarize the data related with the structural features of interaction between the human repair enzyme 8-oxoguanine DNA glycosylase (hOGG1) and DNA. The review covers the questions concerning the role of individual amino acids of hOGG1 in the specific recognition of the oxidized DNA bases, formation of the enzyme-substrate complex, and excision of the lesion bases from DNA. Attention is also focused upon conformational changes in the enzyme active site and disruption of enzyme activity as a result of amino acid mutations. The mechanism of damaged bases release from DNA induced by hOGG1 is discussed in the context of structural dynamics.


Sign in / Sign up

Export Citation Format

Share Document