Reaction of cacodylic acid with organic thiols

Jacobson, K. Bruce; Murphy, Joe; Sarma, B. Das

doi:10.1016/0014-5793(72)80224-2

Cited by 20 publications

(11 citation statements)

References 3 publications

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“…Protomer D has a disordered gap from aa 216-220 in another surface loop of the phosphatase domain. The electron density maps revealed extra density appended to the Cys184 sulfur of protomers A, B, C and D, consistent with a DTT-mediated reaction of cysteine with the arsenic center of the cacodylate (dimethyl arsinic acid) present in the crystallization buffer (Jacobson et al, 1972;Tsao and Maki, 1991;Maignan et al, 1998;Tete-Favier et al, 2000;Junop et al, 2000). The arsenic adduct is located 3 to 3.5 Å above the planar aromatic Trp213 side chain; a similar reaction of a tryptophan-proximal cysteine with cacodylate has been reported for EcoRI methyltransferase and E. coli peptide methionine sulphoxide reductase (Tsao and Maki, 1991;Tete-Favier et al, 2000).…”

Section: New Crystal Structure Of the Pnkp Tetramermentioning

confidence: 61%

Structure–function analysis of the 3′ phosphatase component of T4 polynucleotide kinase/phosphatase

et al. 2007

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T4 polynucleotide kinase/phosphatase (Pnkp) exemplifies a family of bifunctional enzymes with 5'-kinase and 3' phosphatase activities that function in nucleic acid repair. T4 Pnkp is a homotetramer of a 301-aa polypeptide, which consists of an N-terminal kinase domain of the P-loop phosphotransferase superfamily and a C-terminal phosphatase domain of the DxD acylphosphatase superfamily. The homotetramer is formed via pairs of phosphatase-phosphatase and kinase-kinase homodimer interfaces. Here we identify four side chains-Asp187, Ser211, Lys258, and Asp277-that are required for 3' phosphatase activity. Alanine mutations at these positions abolished phosphatase activity without affecting kinase function or tetramerization. Conservative substitutions of asparagine or glutamate for Asp187 did not revive the 3' phosphatase, nor did arginine or glutamine substitutions for Lys258. Threonine in lieu of Ser211 and glutamate in lieu of Asp277 restored full activity, whereas asparagine at position 277 had no salutary effect. We report a 3.0 A crystal structure of the Pnkp tetramer, in which a sulfate ion is coordinated between Arg246 and Arg279 in a position that we propose mimics one of the penultimate phosphodiesters (5'NpNpNp-3') of the polynucleotide 3'-PO(4) substrate. The amalgam of mutational and structural data engenders a plausible catalytic mechanism for the phosphatase that includes covalent catalysis (via Asp165), general acid-base catalysis (via Asp167), metal coordination (by Asp165, Asp277 and Asp278), and transition state stabilization (via Lys258, Ser211, backbone amides, and the divalent cation). Other critical side chains play architectural roles (Arg176, Asp187, Arg213, Asp254). To probe the role of oligomerization in phosphatase function, we introduced six double-alanine cluster mutations at the phosphatase-phosphatase domain interface, two of which (R297A-Q295A and E292A-D300A) converted Pnkp from a tetramer to a dimer and ablated phosphatase activity.

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Section: New Crystal Structure Of the Pnkp Tetramermentioning

confidence: 61%

Structure–function analysis of the 3′ phosphatase component of T4 polynucleotide kinase/phosphatase

et al. 2007

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“…Under acidic conditions, cacodylate can act as an oxidizing agent. The reaction of cacodylate with organic thiols is accompanied by the disappearance of the acid function of cacodylate and the SH groups [26]. The reaction between an aqueous solution of dimethlyarsinic acid and cysteine [(CH 3 ) 2 As(O)OH + 3Cys → (CH 3 ) 2 As‐Cys + cystine + H 2 O] has previously been described [27].…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, interactions between cacodylate and other enzymes have previously been observed [28]. For example, the inhibition by 2‐mercaptoethanol of phenylalanyl‐tRNA synthetase of Neurospora crassa with valine tRNA of Escherichia coli was much more severe in the presence of cacodylate buffer than Tris [26,29]. This is probably due to the formation of a reactive product of cacodylate and mercaptoethanol.…”

Section: Discussionmentioning

confidence: 99%

Inactivation of Aeromonas hydrophila metallo‐β‐lactamase by cephamycins and moxalactam

Zervosen

Hernandez-Valladares

Devreese³

et al. 2001

European Journal of Biochemistry

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Incubation of moxalactam and cefoxitin with the Aeromonas hydrophila metallo‐β‐lactamase CphA leads to enzyme‐catalyzed hydrolysis of both compounds and to irreversible inactivation of the enzyme by the reaction products. As shown by electrospray mass spectrometry, the inactivation of CphA by cefoxitin and moxalactam is accompanied by the formation of stable adducts with mass increases of 445 and 111 Da, respectively. The single thiol group of the inactivated enzyme is no longer titrable, and dithiothreitol treatment of the complexes partially restores the catalytic activity. The mechanism of inactivation by moxalactam was studied in detail. Hydrolysis of moxalactam is followed by elimination of the 3′ leaving group (5‐mercapto‐1‐methyltetrazole), which forms a disulfide bond with the cysteine residue of CphA located in the active site. Interestingly, this reaction is catalyzed by cacodylate.

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“…Crystals of the wild-type a 1 PAS domain (residues 279-404) were initially small and could not be improved, possibly due to a requirement for cysteine modification by the arsenic in the cacodylate-containing crystallization buffer. [21][22][23] To overcome this, we made the triple cysteine mutant C285A, C352A, C374A. This protein crystallized under new conditions, yielding larger crystals with a rhombic dodecahedron morphology and diffraction to 1.8 Å resolution (Table I).…”

Section: Crystal Structure Of the A 1 Pas Domainmentioning

confidence: 99%

Crystal structure of the Alpha subunit PAS domain from soluble guanylyl cyclase

2013

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Soluble guanylate cyclase (sGC) is a heterodimeric heme protein of~150 kDa and the primary nitric oxide receptor. Binding of NO stimulates cyclase activity, leading to regulation of cardiovascular physiology and providing attractive opportunities for drug discovery. How sGC is stimulated and where candidate drugs bind remains unknown. The a and b sGC chains are each composed of Heme-Nitric Oxide Oxygen (H-NOX), Per-ARNT-Sim (PAS), coiled-coil and cyclase domains. Here, we present the crystal structure of the a 1 PAS domain to 1.8 Å resolution. The structure reveals the binding surfaces of importance to heterodimer function, particularly with respect to regulating NO binding to heme in the b 1 H-NOX domain. It also reveals a small internal cavity that may serve to bind ligands or participate in signal transduction.

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Reaction of cacodylic acid with organic thiols

Cited by 20 publications

References 3 publications

Structure–function analysis of the 3′ phosphatase component of T4 polynucleotide kinase/phosphatase

Structure–function analysis of the 3′ phosphatase component of T4 polynucleotide kinase/phosphatase

Inactivation of Aeromonas hydrophila metallo‐β‐lactamase by cephamycins and moxalactam

Crystal structure of the Alpha subunit PAS domain from soluble guanylyl cyclase

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