How is the distal pocket of a heme protein optimized for binding of tryptophan?

Chauhan, Nishma; Basran, Jaswir; Rafice, Sara A.; Efimov, Igor; Millett, Elizabeth S.; Mowat, Christopher G.; Moody, P.C.E.; Handa, Sandeep; Raven, Emma Lloyd

doi:10.1111/febs.12036

Cited by 21 publications

(24 citation statements)

References 32 publications

(55 reference statements)

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“…According to a series of studies employing human IDO1 variants, Ser167 is not regarded to be essential for substrate binding . Indeed, S167A and S167H variants of human IDO1 showed high affinity for l ‐Trp (low K m for l ‐Trp: 21 μ m ) .…”

Section: Resultsmentioning

confidence: 99%

“…According to a series of studies employing human IDO1 variants, Ser167 is not regarded to be essential for substrate binding [17,18,31]. Indeed, S167A and S167H variants of human IDO1 showed high affinity for L-Trp (low K m for L-Trp: 21 lM) [31]. Even in the present study, mouse and platypus IDO1 S171T variants still retain a rather high affinity for L-Trp, although their K m values are 2.8-to 4.5-fold higher than that of wild-types.…”

Section: Evolution Of Vertebrate Idosmentioning

confidence: 99%

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Low efficiency IDO2 enzymes are conserved in lower vertebrates, whereas higher efficiency IDO1 enzymes are dispensable

2015

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Indoleamine 2,3-dioxygenase (IDO) is a Trp-degrading enzyme that catalyzes the first step in the kynurenine pathway. Two IDO genes, IDO1 and IDO2, are found in vertebrates and the timing of the gene duplication giving rise to the genes has been controversial. In the present study, we report that several fishes and two turtles also have both IDO1 and IDO2. This represents definitive evidence for the gene duplication occurring before the divergence of vertebrates, with IDO1 having been lost in a number of lower vertebrate lineages. IDO2 enzymes have a relatively low affinity for L-Trp; however, Anolis carolinensis (lizard) IDO2 has an affinity for L-Trp comparable to mammalian IDO1 enzymes. We identified a Ser residue located in the distal heme pocket of IDO1 (distal-Ser) (corresponding to Ser167 of human IDO1) that is conserved in all IDO1 enzymes and the lizard IDO2. This residue is conserved as Thr (distal-Thr) in other IDO2 enzymes. Biochemical analyses, using IDO variants with either Ser or Thr substitutions, suggest that the distal-Ser change was crucial for the improvement in affinity for L-Trp in ancient IDO1. The ancestral IDO1 likely had a 'moderate' enzymatic efficiency for L-Trp, clearly higher than IDO2 but lower than mammalian IDO1. The distal-Ser of lizard IDO2 bestows a high affinity for L-Trp, however, this unique IDO2 has a low enzymatic efficiency because of its very low catalytic velocity. Thus, low efficiency IDO2 enzymes have been conserved throughout vertebrate evolution, whereas higher efficiency IDO1 enzymes are dispensable in many lower vertebrate lineages.

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Section: Resultsmentioning

confidence: 99%

Section: Evolution Of Vertebrate Idosmentioning

confidence: 99%

Low efficiency IDO2 enzymes are conserved in lower vertebrates, whereas higher efficiency IDO1 enzymes are dispensable

2015

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“…The determinants of substrate binding in these proteins have been probed in molecule detail using sitedirected mutagenesis, 61 following similar detailed studies in peroxidases. 62 These observations are relevant for this study, since spectroscopy has been used to quantify this binding in the dioxygenases.…”

Section: ■ Discussionmentioning

confidence: 99%

Measurement of Internal Substrate Binding in Dehaloperoxidase–Hemoglobin by Competition with the Heme–Fluoride Binding Equilibrium

Zhao

Moretto

et al. 2015

J. Phys. Chem. B

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The application of fluoride anion as a probe for investigating the internal substrate binding has been developed and applied to dehaloperoxidase-hemoglobin (DHP) from Amphitrite ornata. By applying the fluoride titration strategy using UV-vis spectroscopy, we have studied series of halogenated phenols, other substituted phenols, halogenated indoles, and several natural amino acids that bind internally (and noncovalently) in the distal binding pocket of the heme. This approach has identified 2,4-dibromophenol (2,4-DBP) as the tightest binding substrate discovered thus far, with approximately 20-fold tighter binding affinity than that of 4-bromophenol (4-BP), a known internally binding inhibitor in DHP. Combined with resonance Raman spectroscopy, we have confirmed that competitive binding equilibria exist between fluoride anion and internally bound molecules. We have further investigated the hydrogen bonding network of the active site of DHP that stabilizes the exogenous fluoride ligand. These measurements demonstrate a general method for determination of differences in substrate binding affinity based on detection of a competitive fluoride binding equilibrium. The significance of the binding that 2,4-dibromophenol binds more tightly than any other substrate is evident when the structural and mechanistic data are taken into consideration.

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“…Chauhan et al . reported that a Phe163Ala variant shows a drastic decrease in k cat and that approximately 130‐ and 500‐fold increases in K m are observed in Phe163Ala and Arg231Lys variants, respectively. These results suggested that these three residues are crucial for human IDO1 activity.…”

Section: Introductionmentioning

confidence: 99%

High l‐Trp affinity of indoleamine 2,3‐dioxygenase 1 is attributed to two residues located in the distal heme pocket

Yuasa¹

2016

The FEBS Journal

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Indoleamine 2, 3-dioxygenase (IDO) catalyzes the oxidative cleavage of the pyrrole ring of l-Trp to generate N-formyl-kynurenine. Two IDO genes, IDO1 and IDO2, are found in vertebrates. Mammalian IDO1s are high-affinity, l-Trp-degrading enzymes, whereas IDO2s generally have a relatively low affinity. It has been suggested that the distal-Ser (corresponding to Ser167 of human IDO1) was crucial for improvement in the affinity for l-Trp but this idea was insufficient to explain the high affinity shown by mammalian IDO1. In this study, the amino acid sequences of vertebrate ancestral IDO1 and ancestral IDO2 were inferred, and bacterially expressed ancestral IDOs were characterized. Although the amino acid sequences of the enzymes shared high identity (86%) with each other, they showed distinct enzymatic properties. In analyses of a series of ancestral IDO1/IDO2 chimeric enzymes and their variants, the distal-Tyr (corresponding to Tyr126 of human IDO1) was detected as another and was probably the most crucial residue for high l-Trp affinity. The two amino acid substitutions (distal-Ser to Thr and distal-Tyr to His) drastically decreased the l-Trp affinity and catalytic efficiency of IDO1s. Conversely, two substitutions (distal-Thr to Ser and distal-His to Tyr) were sufficient to bestow IDO1-like high affinity on ancestral and chicken IDO2.

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How is the distal pocket of a heme protein optimized for binding of tryptophan?

Cited by 21 publications

References 32 publications

Low efficiency IDO2 enzymes are conserved in lower vertebrates, whereas higher efficiency IDO1 enzymes are dispensable

Low efficiency IDO2 enzymes are conserved in lower vertebrates, whereas higher efficiency IDO1 enzymes are dispensable

Measurement of Internal Substrate Binding in Dehaloperoxidase–Hemoglobin by Competition with the Heme–Fluoride Binding Equilibrium

High l‐Trp affinity of indoleamine 2,3‐dioxygenase 1 is attributed to two residues located in the distal heme pocket

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