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  E.coli news vol. 6  2004.3.2
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This is a digest of daily PubMed searching of E.coli new finding.

***flash report***

  >>>yddE structure<<<

Crystal structure of E. coli YddE protein reveals a striking homolgy with diaminopimelate epimerase (2003) Grassick A, Sulzenbacher G, Roig-Zamboni V, Campanacci V, Cambillau C, Bourne Y Proteins In press:
http://afmb.cnrs-mrs.fr/article57.html

***this year finding***

  >>>novel anaerobic beta-oxidation pathway<<<

Mol Microbiol. 2003 Feb;47(3):793-805. 

A new Escherichia coli metabolic competency: growth on fatty acids by a novel anaerobic beta-oxidation pathway.
Campbell JW, Morgan-Kiss RM, Cronan JE Jr.
Department of Microbiology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.

Escherichia coli uses fatty acids as a sole carbon and energy source during aerobic growth by means of the enzymes encoded by the fad regulon. We report that this bacterium can also grow on fatty acids under anaerobic conditions provided that a terminal respiratory electron acceptor such as nitrate is available. This anaerobic utilization pathway is distinct from the well-studied aerobic pathway in that (i). it proceeds normally in mutant strains lacking various enzymes of the aerobic pathway; (ii). it functions with fatty acids (octanoate and decanoate) that cannot be used by wild-type E. coli strains under aerobic conditions; and (iii). super-repressor mutants of the fadR regulatory locus that block aerobic growth on fatty acids fail to block the anaerobic pathway. We have identified homologues of the FadA, FadB and FadD proteins required for aerobic fatty acid utilization called YfcY, YfcX and YdiD, respectively, which are involved in anaerobic growth on fatty a

cids. A strong FadR binding site was detected upstream of the yfcY gene consistent with microarray analyses, indicating that yfcYX expression is negatively regulated by FadR under aerobic growth conditions. In contrast, transcriptional regulation of ydiD appears to be independent of FadR, and anaerobic growth on fatty acids is not under FadR control. These three genes are conserved in the available genome sequences of pathogenic E. coli, Shigella and Salmonella strains.

PMID: 12535077

  >>>ybgI structure<<<

BMC Struct Biol. 2003 Sep 30;3(1):7. 

 Crystal structure of Escherichia coli protein ybgI, a toroidal structure with a dinuclear metal site.
Ladner JE, Obmolova G, Teplyakov A, Howard AJ, Khil PP, Camerini-Otero RD, Gilliland GL.
Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute and the National Institute of Standards and Technology, 9600 Gudelsky Drive, Rockville, MD 20850, U,S,A.

BACKGROUND: The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function. RESULTS: The structure of the ybgI protein is a toroid composed of six polypeptide chains forming a trimer of dimers. Each polypeptide chain binds two metal ions on the inside of the toroid. CONCLUSION: The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism. The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

PMID: 14519207

***additional finding***

  >>>ribonucleoside hydrolases<<<

   J Biol Chem. 2001 Jan 12;276(2):884-94. 

The RihA, RihB, and RihC ribonucleoside hydrolases of Escherichia coli. Substrate specificity, gene expression, and regulation.
Petersen C, Moller LB.
Department of Biological Chemistry, Institute of Molecular Biology, University of Copenhagen, Solvgade 83H, DK1307 Copenhagen K, Denmark. carstenpt@mermaid.molbio.ku.dk

Pyrimidine-requiring cdd mutants of Escherichia coli deficient in cytidine deaminase utilize cytidine as a pyrimidine source by an alternative pathway. This has been presumed to involve phosphorylation of cytidine to CMP by cytidine/uridine kinase and subsequent hydrolysis of CMP to cytosine and ribose 5-phosphate by a putative CMP hydrolase. Here we show that cytidine, in cdd strains, is converted directly to cytosine and ribose by a ribonucleoside hydrolase encoded by the previously uncharacterized gene ybeK, which we have renamed rihA. The RihA enzyme is homologous to the products of two unlinked genes, yeiK and yaaF, which have been renamed rihB and rihC, respectively. The RihB enzyme was shown to be a pyrimidine-specific ribonucleoside hydrolase like RihA, whereas RihC hydrolyzed both pyrimidine and purine ribonucleosides. The physiological function of the ribonucleoside hydrolases in wild-type E. coli strains is enigmatic, as their activities are paralleled b

y the phosphorolytic activities of the nucleoside phosphorylases, and a triple mutant lacking all three hydrolytic activities grew normally. Furthermore, enzyme assays and lacZ gene fusion analysis indicated that rihB was essentially silent unless activated by mutation, whereas rihA and rihC were poorly expressed in glucose medium due to catabolite repression.

PMID: 11027694

  >>>JW2327, yfcB/prmB, N(5)-glutamine methyltransferase<<<

EMBO J. 2002 Feb 15;21(4):769-78. 

The hemK gene in Escherichia coli encodes the N(5)-glutamine methyltransferase that modifies peptide release factors.
Heurgue-Hamard V, Champ S, Engstrom A, Ehrenberg M, Buckingham RH.
UPR9073 du CNRS, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, Paris 75005, France.

Class 1 peptide release factors (RFs) in Escherichia coli are N(5)-methylated on the glutamine residue of the universally conserved GGQ motif. One other protein alone has been shown to contain N(5)-methylglutamine: E.coli ribosomal protein L3. We identify the L3 methyltransferase as YfcB and show that it methylates ribosomes from a yfcB strain in vitro, but not RF1 or RF2. HemK, a close orthologue of YfcB, is shown to methylate RF1 and RF2 in vitro. hemK is immediately downstream of and co-expressed with prfA. Its deletion in E.coli K12 leads to very poor growth on rich media and abolishes methylation of RF1. The activity of unmethylated RF2 from K12 strains is extremely low due to the cumulative effects of threonine at position 246, in place of alanine or serine present in all other bacterial RFs, and the lack of N(5)-methylation of Gln252. Fast-growing spontaneous revertants in hemK K12 strains contain the mutations Thr246Ala or Thr246Ser in RF2. HemK and YfcB ar

e the first identified methyltransferases modifying glutamine, and are widely distributed in nature.

PMID: 11847124

  >>>group II intron<<<

RNA. 2002 Oct;8(10):1294-307.

The dispersal of five group II introns among natural populations of Escherichia coli.
Dai L, Zimmerly S.
Department of Biological Sciences, University of Calgary, Alberta, Canada.

Group II introns are self-splicing RNAs that also act as retroelements in bacteria, mitochondria, and chloroplasts. Group II introns were identified in Escherichia coli in 1994, but have not been characterized since, and, instead, other bacterial group II introns have been studied for splicing and mobility properties. Despite their apparent intractability, at least five distinct group II introns exist naturally in E. coli strains. To illuminate their function and learn how the introns have dispersed in their natural host, we have investigated their distribution in the ECOR reference collection. Two introns were cloned and sequenced to complete their partial sequences. Unexpectedly, southern blots showed all ECOR strains to contain fragments and/or full-length copies of group II introns, with some strains containing up to 15 intron copies. One intron, E.c.14, has two natural homing sites in IS629 and IS911 elements, and the intron can be present in one, both, or nei

ther homing site in a given strain. Nearly all strains that contain full-length introns also contain unfilled homing sites, suggesting either that mobility is highly inefficient or that most full-length copies are nonfunctional. The data indicate independent mobility of the introns, as well as mobility via the host DNA elements, and overall, the pattern of intron distribution resembles that of IS elements.

PMID: 12403467

*E.coli K-12 W3110 strain contained 2 IS911

  >>>GTPase YjeQ<<<

Biochemistry. 2002 Sep 17;41(37):11109-17. 

YjeQ, an essential, conserved, uncharacterized protein from Escherichia coli, is an unusual GTPase with circularly permuted G-motifs and marked burst kinetics.
Daigle DM, Rossi L, Berghuis AM, Aravind L, Koonin EV, Brown ED.
Antimicrobial Research Centre, Department of Biochemistry, McMaster University, Hamilton, Ontario, Canada L8N 3Z5.

The Escherichia coli protein YjeQ represents a protein family whose members are broadly conserved in bacteria and have been shown to be indispensable to the growth of E. coli and Bacillus subtilis [Arigoni, F., et al. (1998) Nat. Biotechnol. 16, 851]. Proteins of the YjeQ family contain all sequence motifs typical of the vast class of P-loop-containing GTPases, but show a circular permutation, with a G4-G1-G3 pattern of motifs as opposed to the regular G1-G3-G4 pattern seen in most GTPases. All YjeQ family proteins display a unique domain architecture, which includes a predicted N-terminal OB-fold RNA-binding domain, the central permuted GTPase module, and a zinc knuckle-like C-terminal cysteine cluster. This domain architecture suggests a possible role for YjeQ as a regulator of translation. YjeQ was overexpressed, purified to homogeneity, and shown to contain 0.6 equiv of GDP. Steady state kinetic analyses indicated slow GTP hydrolysis, with a k(cat) of 9.4 h(-)(

1) and a K(m) for GTP of 120 microM (k(cat)/K(m) = 21.7 M(-)(1) s(-)(1)). YjeQ also hydrolyzed other nucleoside triphosphates and deoxynucleotide triphosphates such as ATP, ITP, and CTP with specificity constants (k(cat)/K(m)) ranging from 0.2 to 1.0 M(-)(1) s(-)(1). Pre-steady state kinetic analysis of YjeQ revealed a burst of nucleotide hydrolysis for GTP described by a first-order rate constant of 100 s(-)(1) as compared to a burst rate of 0.2 s(-)(1) for ATP. In addition, a variant in the G1 motif of YjeQ (S221A) was substantially impaired for GTP hydrolysis (0.3 s(-)(1)) with a less significant impact on the steady state rate (1.8 h(-)(1)). In summary, E. coli YjeQ is an unusual, circularly permuted P-loop-containing GTPase, which catalyzes GTP hydrolysis at a rate 45 000 times greater than that of turnover.

PMID: 12220175