Proton transfer in the catalytic mechanism of carbonic anhydrase. Effects of placing histidine residues at various positions in the active site of human isoenzyme II

Liang, Zhiwei; Jonsson, Bengt‐Harald; Lindskog, Sven

doi:10.1016/0167-4838(93)90048-v

Cited by 25 publications

(25 citation statements)

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“…The loop residues Glu88 and Glu89 which have been shown to operate the proton transfer minimally in Zn-Cam and the corresponding loop are not found in the Zn-Cap structure. Interestingly, in the case of human carbonic anhydrase it has been observed that the similar process of proton transfer is carried out by His64 (Liang et al, 1993).…”

Section: Active-site Architecturementioning

confidence: 92%

Observation of a calcium-binding site in the γ-class carbonic anhydrase fromPyrococcus horikoshii

Jeyakanthan

Rangarajan

Mridula

et al. 2008

Acta Crystallogr D Biol Crystallogr

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Carbonic anhydrases are zinc-containing metalloenzymes that catalyze the interconversion of carbon dioxide and bicarbonate. Three crystal structures of gamma-class carbonic anhydrase (one of which is bound to a bicarbonate molecule) from the aerobic OT3 strain of the hyperthermophilic archeon Pyrococcus horikoshii have been solved by molecular replacement in space group F4(1)32. The asymmetric unit contains a monomer of 173 amino acids and a catalytic Zn2+ ion. The protein fold is a regular prism formed by a left-handed beta-helix, similar to previously reported structures. The active-site Zn2+ ion located at the interface between the two monomers is bound to three histidyl residues and a water molecule in a tetrahedral fashion. In addition to the 20 beta-strands comprising the beta-helix, there is also a long C-terminal alpha-helix. For the first time, Ca2+ ions have been observed in addition to the catalytic Zn2+ ion. It is hypothesized that Tyr159 (which corresponds to the catalytically important Asn202 in previously reported structures) utilizes C-H...pi interactions to fulfill its functions. This study may shed light on the catalytic mechanism of the enzyme and throw open new questions on the mechanism of product removal in carbonic anhydrases.

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Section: Active-site Architecturementioning

confidence: 92%

Observation of a calcium-binding site in the γ-class carbonic anhydrase fromPyrococcus horikoshii

Jeyakanthan

Rangarajan

Mridula

et al. 2008

Acta Crystallogr D Biol Crystallogr

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“…These experiments use histidine placed at various sites (positions 62, 64, 67, 200) in the active-site cavity and test their capacity to promote proton transfer, following the example of Liang et al [18]. Other experiments use 4-methylimidazole (4-MI) in chemical rescue of mutants lacking a proton shuttle residue, with accompanying crystallographic studies to estimate the binding site of the 4-MI.…”

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confidence: 99%

Proton transfer in catalysis and the role of proton shuttles in carbonic anhydrase

Mikulski

Silverman

2010

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

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The undisputed role of His64 in proton transfer during catalysis by carbonic anhydrases in the α class has raised questions concerning the details of its mechanism. The highly conserved residues Tyr7, Asn62, and Asn67 in the active-site cavity function to fine tune the properties of proton transfer by human carbonic anhydrase II (HCA II). For example, hydrophobic residues at these positions favor an inward orientation of His64 and a low pK a for its imidazole side chain. It appears that the predominant manner in which this fine tuning is achieved in rate constants for proton transfer is through the difference in pK a between His64 and the zinc-bound solvent molecule. Other properties of the active-site cavity, such as inward and outward conformers of His64, appear associated with the change in ΔpK a ; however, there is no strong evidence to date that the inward and outward orientations of His64 are in themselves requirements for facile proton transfer in carbonic anhydrase. Keywordscarbonic anhydrase; proton transfer; carbon dioxide; bicarbonate; hydration An important advance in understanding catalysis by carbonic anhydrase came in the report in 1975 from Steiner, Jonsson, and Lindskog [1] who used hydrogen/deuterium isotope effects to deduce a rate-limiting, intramolecular proton transfer in the maximum velocity of catalysis by human carbonic anhydrase II (HCA II). They suggested that His64 was the likely shuttle group based on its position in the active site and its pK a near 7 that was consistent with the kinetic pK a of k cat . Direct evidence that this suggestion was correct came 14 years later; it was the reduction in maximal velocity by about a factor of 20 caused by the replacement of His64 in HCA II with Ala which cannot support proton transfer [2]. Moreover, maximal velocity catalyzed by the mutant H64A HCA II was rescued to levels close to that of the wild-type enzyme by imidazole buffer in solution [2].Because of its rate-limiting, intramolecular proton transfer in catalysis, carbonic anhydrase has become a model for the examination of long range proton transfers and the role of hydrogenbonded water networks [1,3]. This is a step apparently shared by the numerous carbonic anhydrases classified into genetically distinct classes, such as α, β, and γ [4,5], all of which carry out catalysis of CO 2 hydration in a two-step mechanism. The first stage in the hydration Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access Author ManuscriptBiochim Biophys Acta. Author manuscript; available in PMC 2011 February 1. direction is the reaction of zinc-b...

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“…It has been shown that imidazole derivatives, such as 1,2-dimethylimidazole, provide a relatively efficient 'shunt' pathway for proton transfer without participation of the shuttle group, while other buffers, such as Taps, do not, probably because they cannot enter the active site cavity deeply enough to establish an effective 'shunt' (Tu et al, 1989). In wild-type HCA 11, proton shuttling via His64 is the dominating pathway and the buffer specificity ratio is approximately 1 (Table l), whereas the mutant [H64AH]CA 11, which cannot shuttle protons, has a buffer specificity ratio of 0.03 (Liang et al, 1993). In this mutant, proton transfer in Taps buffer probably occurs between the metal ion center and bulk water molecules.…”

Section: Discussionmentioning

confidence: 99%

Catalytic and Inhibitor‐Binding Properties of Some Active‐Site Mutants of Human Carbonic Anhydrase I

Engstrand

Jonsson

Lindskog

1995

European Journal of Biochemistry

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Three isozyme-specific residues in the active site of human carbonic anhydrase I, Va162, His67, and His200, have been changed by site-directed mutagenesis to their counterparts in human carbonic anhydrase 11, Asn62, Asn67, and Thr200. A double mutant, containing Asn62 and Asn67, and a triple mutant, containing all three alterations, were also produced. The rates of CO, hydration and ester hydrolysis catalyzed by these mutants, the inhibition of these enzymes by the anions, SCN-, and I-, and the binding of the sulfonamide inhibitors, dansylamide and MK-417 (a thienothiopyran-2-sulfonamide) have been measured. The results suggest that the effect of His200 in isozyme I is to prolong the lifetime of the enzyme-bicarbonate complex and to increase the pK, of the catalytic group, a zinc-coordinated water molecule. For isozyme I, Val62 and His67 might interfere with the function of a proton 'shuttle' group in the active site, thus maintaining the buffer specificity of a compulsory proton-transfer step. The single mutations have small effects on anion binding. Only the triple mutant has anion-binding properties resembling those of isozyme 11. All mutants show altered sulfonamide-binding properties. In particular, the binding specificity is affected. While wild-type isozyme I binds dansylamide SO times more strongly than MK-417, the triple mutant shows a reversed selectivity and binds MK-417 nearly SO times more strongly than dansylamide.

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Proton transfer in the catalytic mechanism of carbonic anhydrase. Effects of placing histidine residues at various positions in the active site of human isoenzyme II

Cited by 25 publications

References 20 publications

Observation of a calcium-binding site in the γ-class carbonic anhydrase fromPyrococcus horikoshii

Observation of a calcium-binding site in the γ-class carbonic anhydrase fromPyrococcus horikoshii

Proton transfer in catalysis and the role of proton shuttles in carbonic anhydrase

Catalytic and Inhibitor‐Binding Properties of Some Active‐Site Mutants of Human Carbonic Anhydrase I

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