Related Articles31P NMR and genetic analysis establish hinT as the only Escherchia coli purine nucleoside phosphoramidase and as essential for growth under high salt conditions.
Eukaryotic cells encode AMP-lysine (AMP-N-epsilon-(N-alpha-acetyl lysine methyl ester) 5'-phosphoramidate) hydrolases related to the rabbit histidine triad nucleotide-binding protein 1 (Hint1) sequence. Bacterial and archaeal cells have Hint homologs annotated in a variety of ways, but the enzymes have not been characterized, nor have phenotypes been described due to loss of enzymatic activity. We developed a quantitative (31)P NMR assay to determine whether Escherichia coli possesses an adenosine phosphoramidase activity. Indeed, soluble lysates prepared from wild-type laboratory E. coli exhibited activity on the model substrate adenosine 5'-monophosphoramidate (AMP-NH(2)). The E. coli Hint homolog, which had been comprehensively designated ycfF and is here named hinT, was cloned, overexpressed, purified, and characterized with respect to purine nucleoside phosphoramidate substrates. Bacterial hinT was several times more active than human or rabbit Hint1 on five model substrates. In addition, bacterial and mammalian enzymes preferred guanosine versus adenosine phosphoramidates as substrates. Analysis of the lysates from a constructed hinT knock-out strain of E. coli demonstrated that all of the cellular purine nucleoside phosphoramidase activity is due to hinT. Physiological analysis of this mutant revealed that the loss of hinT results in failure to grow in media containing 0.75 m KCl, 0.9 m NaCl, 0.5 m NaOAc, or 10 mm MnCl(2). Thus, cation-resistant bacterial cell growth may be dependent on the hydrolysis of adenylylated and/or guanylylated phosphoramidate substrates by hinT.
Structure analysis of membrane-reconstituted subunit c-ring of E. coli H+-ATP synthase by solid-state NMR.
Structure analysis of membrane-reconstituted subunit c-ring of E. coli H+-ATP synthase by solid-state NMR.
Structure analysis of membrane-reconstituted subunit c-ring of E. coli H+-ATP synthase by solid-state NMR.
J Biomol NMR. 2010 Sep;48(1):1-11
Authors: Todokoro Y, Kobayashi M, Sato T, Kawakami T, Yumen I, Aimoto S, Fujiwara T, Akutsu H
The subunit c-ring of H(+)-ATP synthase (F(o) c-ring) plays an essential role in the proton translocation across a membrane driven by the electrochemical potential. To understand its structure and function, we...
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12-18-2010 12:00 PM
NMR analysis reveals 17?-estradiol induced conformational change in ER? ligand binding domain expressed in E. coli.
NMR analysis reveals 17?-estradiol induced conformational change in ER? ligand binding domain expressed in E. coli.
NMR analysis reveals 17?-estradiol induced conformational change in ER? ligand binding domain expressed in E. coli.
Mol Biol Rep. 2010 Dec 12;
Authors: Paramanik V, Thakur MK
Nuclear magnetic resonance (NMR) spectroscopy is a useful biophysical technique to study the ligand-protein interaction. In this report, we have used bacterially produced ER? and its domains for studying the functional analysis of ligand-protein interaction....
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12-15-2010 12:03 PM
[NMR paper] Resonance assignments and secondary structure analysis of E. coli thioredoxin by magic angle spinning solid-state NMR spectroscopy.
Resonance assignments and secondary structure analysis of E. coli thioredoxin by magic angle spinning solid-state NMR spectroscopy.
Related Articles Resonance assignments and secondary structure analysis of E. coli thioredoxin by magic angle spinning solid-state NMR spectroscopy.
J Phys Chem B. 2005 Sep 29;109(38):18135-45
Authors: Marulanda D, Tasayco ML, Cataldi M, Arriaran V, Polenova T
De novo site-specific 13C and 15N backbone and sidechain resonance assignments are presented for uniformly enriched E. coli thioredoxin, established using...
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[NMR paper] GANA--a genetic algorithm for NMR backbone resonance assignment.
GANA--a genetic algorithm for NMR backbone resonance assignment.
Related Articles GANA--a genetic algorithm for NMR backbone resonance assignment.
Nucleic Acids Res. 2005;33(14):4593-601
Authors: Lin HN, Wu KP, Chang JM, Sung TY, Hsu WL
NMR data from different experiments often contain errors; thus, automated backbone resonance assignment is a very challenging issue. In this paper, we present a method called GANA that uses a genetic algorithm to automatically perform backbone resonance assignment with a high degree of precision and recall....
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[NMR paper] NMR analysis of cleaved Escherichia coli thioredoxin (1-73/74-108) and its P76A varia
NMR analysis of cleaved Escherichia coli thioredoxin (1-73/74-108) and its P76A variant: cis/trans peptide isomerization.
Related Articles NMR analysis of cleaved Escherichia coli thioredoxin (1-73/74-108) and its P76A variant: cis/trans peptide isomerization.
Protein Sci. 2000 Jan;9(1):20-8
Authors: Yu WF, Tung CS, Wang H, Tasayco ML
Inspection of high resolution three-dimensional (3D) structures from the protein database reveals an increasing number of cis-Xaa-Pro and cis-Xaa-Yaa peptide bonds. However, we are still far from being able to...
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[NMR paper] Sequential assignment of 2D-NMR spectra of proteins using genetic algorithms.
Sequential assignment of 2D-NMR spectra of proteins using genetic algorithms.
Related Articles Sequential assignment of 2D-NMR spectra of proteins using genetic algorithms.
J Chem Inf Comput Sci. 1993 Mar-Apr;33(2):245-51
Authors: Wehrens R, Lucasius C, Buydens L, Kateman G
The application of genetic algorithms to the problem of the sequential assignment of two-dimensional protein NMR spectra is discussed. The problem is heavily underconstrained since in most cases more patterns are available than amino acid positions, and uncertainties may...
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[NMR paper] NMR study of the phosphoryl binding loop in purine nucleotide proteins: evidence for
NMR study of the phosphoryl binding loop in purine nucleotide proteins: evidence for strong hydrogen bonding in human N-ras p21.
Related Articles NMR study of the phosphoryl binding loop in purine nucleotide proteins: evidence for strong hydrogen bonding in human N-ras p21.
Biochemistry. 1990 Apr 10;29(14):3509-14
Authors: Redfield AG, Papastavros MZ
The structure of the phosphoryl binding region of human N-ras p21 was probed by using heteronuclear proton-observed NMR methods. Normal protein and a Gly-12----Asp-12 mutant protein were prepared...
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Structure analysis of membrane-reconstituted subunit c-ring of E. coli H+-ATP synthas
Abstract The subunit c-ring of H+-ATP synthase (Fo c-ring) plays an essential role in the proton translocation across a membrane driven by the electrochemical potential. To understand its structure and function, we have carried out solid-state NMR analysis under magic-angle sample spinning. The uniformly -labeled Fo c from E. coli (EFo c) was reconstituted into lipid membranes as oligomers. Its high resolution two- and three-dimensional spectra were obtained, and the 13C and 15N signals were assigned. The obtained chemical shifts suggested that EFo c takes on a hairpin-type helix-loop-helix...
31P NMR and genetic analysis establish hinT as the only Escherchia coli purine nucleo
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