Creatine kinase. Nuclear magnetic resonance and fluorescence evidence for interaction of adenosine 5'-diphosphate with aromatic residue(s)

Vašák, Milan; Nagayama, Kuniaki; Wüthrich, Kurt; Ml, Mertens; Jh, Kägi

doi:10.1021/bi00590a004

Cited by 63 publications

(34 citation statements)

References 26 publications

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“…CK is likely to be a 2-domain hinge-bending enzyme like the sequence-related ArgK (Dumas & Janin, 1983), implying that the nucleotide binding domain should be mainly constituted by the C-terminal half of the primary sequence; this is also the case for 3-phosphoglycerate kinase (Watson et al, 1982), which shows some sequence homologies with CK in the C-terminal part. The enzyme kinetic data of our Trp-223 mutants indicate that the active-site tryptophan residue does not directly participate in adenine substrate binding, although previous fluorescence spectroscopic observations suggest a very close vicinity of the indole group and the adenine substrate (Vasak et al, 1979;Messmer & Kagi, 1985). However, these apparently conflicting findings can be brought to agreement by assuming that Trp-223 might be moved into a position close to the adenine ring by a conformational change (e.g., a cleft-closing) after the nucleotide has already bound.…”

Section: Trp-223contrasting

confidence: 67%

“…In CK, an unidentified tryptophan residue has been previously shown to be located at the active site (Vasak et al, 1979;Zhou & Tsou, 1985); in Mib-CK, another tryptophan residue has been postulated to be situated at the interface between the dimers constituting an octamer (Gross & Wallimann, 1993). In the present study, all 5 individual indole side chains of Mib-CK have been replaced by cysteine and/or phenylalanine residues using sitedirected mutagenesis, in order to identify the sequence positions of these 2 functionally defined residues.…”

Section: Discussionmentioning

confidence: 90%

“…Previous spectroscopic studies (Vasak et al, 1979;Messmer & Kagi, 1985) strongly suggest that the active-site tryptophan is part of the adenine binding site of CK. The conservation of Trp-223 in ArgK and CK supports this assumption because both guanidino kinases have the adenine nucleotide substrate in common, whereas the guanidino substrates differ.…”

Section: Trp-223mentioning

confidence: 96%

“…In addition, an involvement of indole side chains in enzymatic activity could be demonstrated by the inactivation of the enzyme by selective chemical modification of 1 tryptophan per protomer (Zhou & Tsou, 1985). Strong evidence for the proximity of a n indole group to the bound adenine substrate has been provided by NMR and fluorescence spectroscopy (Vasak et al, 1979), as well as by circular dichroism studies (Kagi et al, 1971). …”

mentioning

confidence: 99%

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The tryptophan residues of mitochondrial creatine kinase: Roles of Trp‐223, Trp‐206, and Trp‐264 in active‐site and quaternary structure formation

et al. 1994

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The 5 tryptophan residues of chicken sarcomeric mitochondrial creatine kinase (Mi,-CK) were individually replaced by phenylalanine or cysteine using site-directed mutagenesis. The mutant proteins were analyzed by enzyme kinetics, fluorescence spectroscopy, circular dichroism, and conformational stability studies. In the present work, Trp-223 is identified as an active-site residue whose replacement even by phenylalanine resulted in 296% inactivation of the enzyme. Trp-223 is responsible for a strong (18-21%) fluorescence quenching effect occurring upon formation of a transition state-analogue complex (TSAC; Mib-CK .creatine.MgADP .NO,-), and Trp-223 is probably required for the conformational change leading to the TSAC-induced octamer dissociation of Mib-CK.Replacement of Trp-206 by cysteine led to a destabilization of the active-site structure, solvent exposure of Trp-223, and to the dissociation of the Mib-CK dimers into monomers. However, this dimer dissociation was counteracted by TSAC formation or the presence of ADP alone. Trp-264 is shown to be located at the dimerdimer interfaces within the Mib-CK octamer, being the origin of another strong (25%) fluorescence quenching effect, which was observed upon the TSAC-induced octamer dissociation. Substitution of Trp-264 by cysteine drastically accelerated the TSAC-induced dissociation and destabilized the octameric structure by one-fourth of the total free interaction energy, probably by weakening hydrophobic contacts. The roles of the other 2 tryptophan residues, Trp-213 and Trp-268, could be less well assigned.

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Section: Trp-223contrasting

confidence: 67%

Section: Discussionmentioning

confidence: 90%

Section: Trp-223mentioning

confidence: 96%

mentioning

confidence: 99%

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The tryptophan residues of mitochondrial creatine kinase: Roles of Trp‐223, Trp‐206, and Trp‐264 in active‐site and quaternary structure formation

et al. 1994

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“…First, quenching could be related to the spectral overlap of the base absorption red-edge with the fluorescence spectrum of some Trp residues which produces a Forster energy transfer, as proposed in the case of nucleotide binding to creatine kinase [31] and mitochondria1 F,-ATPase [32]. Second, nucleotide binding could generate a conformational change in the connector, which would bring some Trp residues to a higher-quenching environment.…”

Section: Discussionmentioning

confidence: 99%

An intrinsic‐tryptophan‐fluorescence study of phage φ29 connector/nucleic acid interactions

Urbaneja

Rivas

Carrascosa

et al. 1994

European Journal of Biochemistry

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The protein p10 of bacteriophage @29 assembled into connectors exhibit an intrinsic fluorescence with an emission peak centered at 335 nm, which suggests a hydrophobic environment of the three tryptohan residues that the protein contains. Upon incubation with linear DNA (but not with circular DNA), a decrease in the connector intrinsic fluorescence is measured which does not show any sequence specificity. The decrease in fluorescence is not observed when DNA is incubated with proteolyzed connectors, which lack the DNA-binding domain, suggesting that the fluorescence quenching is related to the binding of DNA to the @29 connectors. Acrylamide quenching studies reveal a higher accessibility of tryptophan residues to the quencher when the connector is bound to DNA. Protein denaturation by guanidine hydrochloride occurs at lower denaturant concentrations in the presence of linear DNA (but not circular DNA) than in its absence, suggesting a conformational change of @29 connector upon binding to linear DNA. This hypothesis is supported by the fact that the proteolyzed connectors, which do not bind DNA, are denatured at the same denaturant concentration, regardless of the presence of DNA. @29 connectors also bind RNA, but this interaction does not exert any effect on acrylamide quenching or guanidine hydrochloride denaturation. This result, together with that showing that proteolyzed connectors are able to interact with RNA, reinforces the idea that @29 connectors have two independent domains for interaction with DNA and RNA.Doubled-stranded DNA bacteriophages hold a viral component called portal structure or connector that is required in several important steps of their morphogenetic pathway, such as the prohead formation [l], DNA packaging into the prohead [2] and tail assembly [3]. Structural analysis of several of these connectors has revealed a common stucture [4]: a 12-fold (or 13-fold) wide domain facing the virion capsid and a narrower 6-fold domain facing the virion tail. A channel runs longitudinally through these two domains and it has been proposed to be the site where DNA translocation takes place.Isolated bacteriophage @29 connectors [5] composed of 12 monomers of a single protein (p10, molecular mass 35800 Da) have been shown to bind both linear [6] and circular dsDNA [7] in a sequence-independent way. In addition, assays performed with hybrid Al@29 proheads assembled on @29 connectors [8] have revealed that the latter could package linear but not circular dsDNA into viral proheads [9].It has been shown that a specific RNA is absolutely required for DNA packaging into @29 [lo], whereas when hybrid iU@29 proheads are used, all the RNAs tested have been able to induce DNA packaging, albeit with less efficiency than the specific RNA [ll]. It has also been shown that RNA binds to the @29 connector in a sequence-independent fashion and that protein p10 contains two different domains for DNA and RNA binding [12]. The protein p10 contains three tryptophan residues in its sequence and due to the high sensitivity...

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Creatine Kinase: Structure‐Activity Relationships

Kenyon

Reed

1983

Advances in Enzymology - And Related Areas of Molecular Biology

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Creatine kinase. Nuclear magnetic resonance and fluorescence evidence for interaction of adenosine 5'-diphosphate with aromatic residue(s)

Cited by 63 publications

References 26 publications

The tryptophan residues of mitochondrial creatine kinase: Roles of Trp‐223, Trp‐206, and Trp‐264 in active‐site and quaternary structure formation

The tryptophan residues of mitochondrial creatine kinase: Roles of Trp‐223, Trp‐206, and Trp‐264 in active‐site and quaternary structure formation

An intrinsic‐tryptophan‐fluorescence study of phage φ29 connector/nucleic acid interactions

Creatine Kinase: Structure‐Activity Relationships

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