Biomedical subjects

T Leisinger

Publications and source records attributed to T Leisinger.

At least 37 records · Page 2Linked to original sources

PubMed1999

Properties of the methylcobalamin:H4folate methyltransferase involved in chloromethane utilization by Methylobacterium sp. strain CM4.

Methylobacterium sp. strain CM4 is a strictly aerobic methylotrophic proteobacterium growing with chloromethane as the sole carbon and energy source. Genetic evidence and measurements of enzyme activity in cell-free extracts have suggested a multistep pathway for the conversion of chloromethane to formate. The postulated pathway is initiated by a corrinoid-dependent methyltransferase system involving methyltransferase I (CmuA) and methyltransferase II (CmuB), which transfer the methyl group of chloromethane onto tetrahydrofolate (H4folate) [Vannelli et al. (1999) Proc. Natl Acad. Sci. USA 96, 4615-4620]. We report the overexpression in Escherichia coli and the purification to apparent homogeneity of methyltransferase II. This homodimeric enzyme, with a subunit molecular mass of 33 kDa, catalyzed the conversion of methylcobalamin and H4folate to cob(I)alamin and methyl-H4folate with a specific activity of 22 nmol x min-1 x (mg protein)-1. The apparent kinetic constants for H4folate were: Km = 240 microM, Vmax = 28.5 nmol x min-1 x (mg protein)-1. The reaction appeared to be first order with respect to methylcobalamin at concentrations up to 2 mM, presumably reflecting the fact that methylcobalamin is an artificial substitute for the methylated methyltransferase I, the natural substrate of the enzyme. Tetrahydromethanopterin, a coenzyme also present in Methylobacterium, did not serve as a methyl group acceptor for methyltransferase II. Purified methyltransferase II restored chloromethane dehalogenation by a cell free extract of a strain CM4 mutant defective in methyltransferase II.

Amino Acid Sequence↗

PubMed1998

A family of flavoproteins in the domains Archaea and Bacteria.

A family of flavoproteins, called A-type flavoproteins, is described. It consists of 14 protein sequences of 385-597 amino acids in length, 7 from methanogens (domain: Archaea), 5 from phototrophic prokaryotes, one from Escherichia coli, and a partial sequence from the sulfate reducer Desulfovibrio gigas (domain: Bacteria). No similar sequence could be found in the domain Eucarya. All sequences show significant similarity over a 385-400 amino acid portion overlapping a recognizable flavodoxin signature starting at positions 245-285 of the common core sequence. Cofactor analysis and, to some extent, analysis of the primary structure of six A-type flavoproteins, three of which are structurally characterized here, support the existence of four sub-families: (a) simple flavoproteins binding only FMN; (b) diflavin flavoproteins binding FMN and FAD; (c) a flavorubredoxin binding FMN and iron; (d) a hemoflavoprotein. The possible involvement of A-type flavoproteins in the metabolism of oxygen, as suggested for D. gigas hemoflavoprotein [Gomes, C. M., Silva, G., Oliveira, S., LeGall, J., Liu, M.-Y., Xavier, A. V., Rodrigues-Pousada, C. & Teixeira, M. (1997) J. Biol. Chem. 272, 22502-22508], is discussed.

Amino Acid Sequence↗

PubMed1998

Chloromethane Metabolism by Methylobacterium sp. Strain CM4

Methylobacterium sp. strain CM4 metabolized chloromethane quantitatively with a molar yield of 2.8 g of whole-cell protein/mol of C. This value was similar to that observed after growth with methanol (2.9 g of protein/mol of C) and about three times larger than the yield with formate (0.94 g of protein/mol of C). Chloromethane dehalogenation activity was inducible. MiniTn5 transposon insertion mutants with altered growth characteristics with chloromethane and other C1 compounds were isolated and characterized. Nine of these were unable to grow with chloromethane but were able to grow with methanol, methylamine, or formate. Seventy-three transposon mutants that were defective in the utilization of either methanol, methylamine, methanol plus methylamine, or formate could still grow with chloromethane. Based on the protein yield data and the properties of the transposon mutants, we propose a pathway for chloromethane metabolism that depends on methyltransferase and dehydrogenase activities.

Journal Article↗

PubMed1998

Molecular analysis of Methanobacterium phage psiM2.

The methanogenic archaeon Methanobacterium thermoautotrophicum Marburg is infected by the double-stranded DNA phage psiM2. The complete phage genome sequence of 26 111 bp was established. Thirty-one open reading frames (orfs), all of them organized in the same direction of transcription, were identified. On the basis of comparison of the deduced amino acid sequences to known proteins and by searching for conserved motifs, putative functions were assigned to the products of six orfs. These included three proteins involved in packaging DNA into the capsid, two putative phage structural proteins and a protein related to the Int family of site-specific recombinases. Analysis of the N-terminal amino acid sequences of three phage-encoded proteins led to the identification of two genes encoding structural proteins and of peiP, the structural gene of pseudomurein endoisopeptidase. This enzyme is involved in the lysis of host cells, and it appears to belong to a novel enzyme family. peiP was overexpressed in Escherichia coli, and its product was shown to catalyse the in vitro lysis of M. thermoautotrophicum cells. Comparison of the phage psiM2 DNA sequence with parts of the sequence of the wild-type phage psiM1 suggests that psiM2 is a deletion derivative, which formed by homologous recombination between two copies of a direct repeat.

Amino Acid Sequence↗

PubMed1998

Metabolism of Dichloromethane by the Strict Anaerobe Dehalobacterium formicoaceticum.

The metabolism of dichloromethane by Dehalobacterium formicoaceticum in cell suspensions and crude cell extracts was investigated. The organism is a strictly anaerobic gram-positive bacterium that utilizes exclusively dichloromethane as a growth substrate and ferments this compound to formate and acetate in a molar ratio of 2:1. When [C]dichloromethane was degraded by cell suspensions, formate, the methyl group of acetate, and minor amounts of methanol were labeled, but there was no nuclear magnetic resonance signal corresponding to the carboxyl group of acetate. This finding and previously established carbon and electron balances suggested that dichloromethane was converted to methylene tetrahydrofolate, of which two-thirds was oxidized to formate while one-third gave rise to acetate by incorporation of CO(2) from the medium in the acetyl coenzyme A synthase reaction. When crude desalted extracts were incubated in the presence of dichloromethane, tetrahydrofolate, ATP, methyl viologen, and molecular hydrogen, dichloromethane and tetrahydrofolate were consumed, with the concomitant formation of stoichiometric amounts of methylene tetrahydrofolate. The in vitro transfer of the methylene group of dichloromethane onto tetrahydrofolate required substoichiometric amounts of ATP. The reaction was inhibited in a light-reversible fashion by 20 muM propyl iodide, thus suggesting involvement of a Co(I) corrinoid in the anoxic dehalogenation of dichloromethane. D. formicoaceticum exhibited normal growth with 0.8 mM sodium in the medium, and crude extracts contained ATPase activity that was partially inhibited by N,N'-dicyclohexylcarbodiimide and azide. During growth with dichloromethane, the organism thus may conserve energy not only by substrate-level phosphorylation but also by a chemiosmotic mechanism involving a sodium-independent F(0)F(1)-type ATP synthase.

Journal Article↗

PubMed1998

Effects of bacterial host and dichloromethane dehalogenase on the competitiveness of methylotrophic bacteria growing with dichloromethane.

Methylobacterium sp. strain DM4 and Methylophilus sp. strain DM11 can grow with dichloromethane (DCM) as the sole source of carbon and energy by virtue of homologous glutathione-dependent DCM dehalogenases with markedly different kinetic properties (the kcat values of the enzymes of these strains are 0.6 and 3.3 S-1, respectively, and the Km values are 9 and 59 microM, respectively). These strains, as well as transconjugant bacteria expressing the DCM dehalogenase gene (dcmA) from DM11 or DM4 on a broad-host-range plasmid in the background of dcmA mutant DM4-2cr, were investigated by growing them under growth-limiting conditions and in the presence of an excess of DCM. The maximal growth rates and maximal levels of dehalogenase for chemostat-adapted bacteria were higher than the maximal growth rates and maximal levels of dehalogenase for batch-grown bacteria. The substrate saturation constant of strain DM4 was much lower than the Km of its associated dehalogenase, suggesting that this strain is adapted to scavenge low concentrations of DCM. Strains and transconjugants expressing the DCM dehalogenase from strain DM11, on the other hand, had higher growth rates than bacteria expressing the homologous dehalogenase from strain DM4. Competition experiments performed with pairs of DCM-degrading strains revealed that a strain expressing the dehalogenase from DM4 had a selective advantage in continuous culture under substrate-limiting conditions, while strains expressing the DM11 dehalogenase were superior in batch culture when there was an excess of substrate. Only DCM-degrading bacteria with a dcmA gene similar to that from strain DM4, however, were obtained in batch enrichment cultures prepared with activated sludge from sewage treatment plants.

Base Sequence↗

PubMed1997

Identification of a novel determinant of glutathione affinity in dichloromethane dehalogenases/glutathione S-transferases.

Bacterial dichloromethane dehalogenases catalyze the glutathione-dependent hydrolysis of dichloromethane to formaldehyde and are members of the enzyme superfamily of glutathione S-transferases involved in the detoxification of electrophilic compounds. Numerous protein engineering studies have addressed questions pertaining to the substrate specificity, the reaction mechanism, and the kinetic pathway of glutathione S-transferases. In contrast, the molecular determinants for binding of the glutathione cofactor have been less well investigated. Dichloromethane dehalogenases from Hyphomicrobium sp. DM2 and Methylobacterium sp. DM4 displayed significantly different affinities for glutathione, but not for the dichloromethane substrate. The sequence of dcmA, the dichloromethane dehalogenase gene from strain DM2, was determined and featured a single base difference from the previously determined sequence of dcmA from strain DM4. This base change resulted in a single amino acid difference in the corresponding proteins at sequence position 27. Site-directed variants of the homologous dichloromethane dehalogenase from Methylophilus sp. DM11 (56% amino acid identity) at the corresponding residue in the protein sequence provided further evidence that this residue selectively modulated the dependence of dichloromethane dehalogenase activity on glutathione.

Amino Acid Sequence↗

PubMed1997

Characterization of alpha-ketoglutarate-dependent taurine dioxygenase from Escherichia coli.

The Escherichia coli tauD gene is required for the utilization of taurine (2-aminoethanesulfonic acid) as a sulfur source and is expressed only under conditions of sulfate starvation. The sequence relatedness of the TauD protein to the alpha-ketoglutarate-dependent 2,4-dichlorophenoxyacetate dioxygenase of Alcaligenes eutrophus suggested that TauD is an alpha-ketoglutarate-dependent dioxygenase catalyzing the oxygenolytic release of sulfite from taurine (van der Ploeg, J. R., Weiss, M. A., Saller, E., Nashimoto, H., Saito, N., Kertesz, M. A., and Leisinger, T. (1996) J. Bacteriol. 178, 5438-5446). TauD was overexpressed in E. coli to approximately 70% of the total soluble protein and purified to apparent homogeneity by a simple two-step procedure. The apparent Mr of 81,000 of the native protein and the subunit Mr of 37,400 were consistent with a homodimeric structure. The pure enzyme converted taurine to sulfite and aminoacetaldehyde, which was identified by high pressure liquid chromatography after enzymatic conversion to ethanolamine. The reaction also consumed equimolar amounts of oxygen and alpha-ketoglutarate; ferrous iron was absolutely required for activity; and ascorbate stimulated the reaction. The properties and amino acid sequence of this enzyme thus define it as a new member of the alpha-ketoglutarate-dependent dioxygenase family. The pure enzyme showed maximal activity at pH 6.9 and retained activity on storage at -20 degrees C for several weeks. Taurine (Km = 55 microM) was the preferred substrate, but pentanesulfonic acid, 3-(N-morpholino)propanesulfonic acid, and 1,3-dioxo-2-isoindolineethanesulfonic acid were also desulfonated at significant rates. Among the cosubstrates tested, only alpha-ketoglutarate (Km = 11 microM) supported significant dioxygenase activity.

Amino Acid Sequence↗

PubMed1997

Association of newly discovered IS elements with the dichloromethane utilization genes of methylotrophic bacteria.

Dichloromethane (DCM) dehalogenases enable facultative methylotrophic bacteria to utilize DCM as sole carbon and energy source. DCM-degrading aerobic methylotrophic bacteria expressing a type A DCM dehalogenase were previously shown to share a conserved 4.2 kb BamHI DNA fragment containing the dehalogenase structural gene, dcmA, and dcmR, the gene encoding a putative regulatory protein. Sequence analysis of a 10 kb DNA fragment including this region led to the identification of three types of insertion sequences identified as IS1354, IS1355 and IS1357, and also two ORFs, orf353 and orf192, of unknown function. Two identical copies of element IS1354 flank the conserved 4.2 kb fragment as a direct repeat. The occurrence of these newly identified IS elements was shown to be limited to DCM-utilizing methylotrophs containing a type A DCM dehalogenase. The organization of the corresponding dcm regions in 12 DCM-utilizing strains was examined by hybridization analysis using IS-specific probes. Six different groups could be defined on the basis of the occurrence, position and copy number of IS sequences. All groups shared a conserved 5.6 kb core region with dcmA, dcmR, orf353 and orf192 as well as IS1357. One group of strains including Pseudomonas sp. DM1 contained two copies of this conserved core region. The high degree of sequence conservation observed within the genomic region responsible for DCM utilization and the occurrence of clusters of insertion sequences in the vicinity of the dcm genes suggest that a transposon is involved in the horizontal transfer of the DCM-utilization character among methylotrophic bacteria.

Amino Acid Sequence↗

PubMed1997

Characterization of Methanobacterium thermoautotrophicum Marburg mutants defective in regulation of L-tryptophan biosynthesis.

Three nitrosoguanidine-induced mutants of the archaeon Methanobacterium thermoautotrophicum Marburg resistant to 5-methyltryptophan were isolated and characterized. They were found to take up L-tryptophan, as wild-type cells, via an energy-dependent, low-affinity transport system specific for L-tryptophan, with a Km of 300 microM and a Vmax of 7 nmol/mg (dry weight)/min. Resistance to 5-methyltryptophan was not due to feedback-resistant anthranilate synthase but to constitutive expression of the trp genes, as measured by the specific activities of anthranilate synthase and tryptophan synthase, the enzymes encoded by trpEG and trpB, respectively, of the trpEGCFBAD gene cluster. Estimation of trpE mRNA obtained from mutant cells grown in minimal medium with or without L-tryptophan suggested that constitutive expression resulted from deficient transcriptional regulation. The enhanced expression of the trp genes in the mutants was found to result in intracellular L-tryptophan pools that were two- to fourfold higher than in the wild type. Sequencing of the region upstream of trpE revealed in two mutants point mutations mapping on the 5'-side of the archaeal box A, whereas in the third mutant this region did not differ from that of the wild type. These results suggest that (i) in M. thermoautotrophicum the 5-methyltryptophan-resistant phenotype arises from lesions in components of a regulatory system controlling transcription of the trp genes and (ii) cis-acting sequence elements in front of the trpE promoter may form part of this system.

Base Sequence↗

PubMed1997

Involvement of CysB and Cbl regulatory proteins in expression of the tauABCD operon and other sulfate starvation-inducible genes in Escherichia coli.

Starvation for sulfate results in increased synthesis of several proteins in Escherichia coli. Among these Ssi (sulfate starvation-induced) proteins are the products of the tauABCD genes, which are required for utilization of taurine as sulfur source for growth. In this study, the role of the cbl gene in expression of tauABCD and other ssi genes was investigated. The protein encoded by cbl shows high sequence similarity to CysB, the LysR-type transcriptional activator of the genes involved in cysteine biosynthesis. Strain EC2541, which contains an internal deletion in cbl, was unable to utilize taurine and other aliphatic sulfonates as sulfur sources. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that many of the Ssi proteins were not synthesized in EC2541. Expression of a translational tauD'-'lacZ fusion required the presence of both cbl and cysB. The interactions of CysB and Cbl with the promoter region of tauABCD were studied by using gel mobility shift experiments and DNase I footprinting. CysB occupied multiple binding sites, whereas Cbl occupied only one site from 112 to 68 bp upstream of the transcription start site. Acetylserine, the inducer of transcription of CysB-regulated genes, stimulated binding of CysB but not of Cbl. Sulfate had no effect on binding of both proteins to the tauABCD promoter region. These results indicate that Cbl is a transcription factor for genes required for sulfonate-sulfur utilization and maybe for other genes whose expression is induced by sulfate starvation.

ATP-Binding Cassette Transporters↗

PubMed1996

Protein engineering studies of dichloromethane dehalogenase/glutathione S-transferase from Methylophilus sp. strain DM11. Ser12 but not Tyr6 is required for enzyme activity.

The structural gene for dichloromethane dehalogenase/glutathione S-transferase (GST, EC 2.5.1.18) from Methylophilus sp. strain DM11 was subcloned into a multicopy plasmid under the control of the T7 polymerase promoter, allowing expression in Escherichia coli and easy purification of the enzyme in good yield. Several point mutations leading to amino acid changes at residues Tyr6, His8 and Ser12 of the protein were introduced in this gene. Mutations at Tyr6, the N-terminal tyrosine known to be essential for enzymatic activity in glutathione S-transferases of the alpha, mu, and pi classes, had little effect on the activity of dichloromethane dehalogenase. The same applied for mutations at residue His8, which from multiple alignments of GST sequences may also correspond to the conserved N-terminal tyrosine residue of GST enzymes. The higher turnover rate of the wild-type enzyme with dibromomethane compared with dichloromethane was lost in mutants with amino acid replacements at residue His8, but retained in mutant proteins at Tyr6. Mutations at Ser12 led to mutants with drastically reduced enzymatic activity, pinpointing this residue as an essential determinant of catalytic efficiency.

Amino Acid Sequence↗

PubMed1996

Biodegradation of chlorinated aliphatic compounds.

During the past year, the range of environmentally relevant chlorinated aliphatic compounds known to serve as growth substrates for pure cultures of bacteria has been extended and novel reactions for the aerobic co-metabolic transformation of chloroaliphatics have been reported. The biochemistry of chloroaliphatics degradation in the new aerobic isolates is still unexplored, but progress has been made in understanding some of the anaerobic dehalogenation reactions.

Bacteria, Aerobic↗

PubMed1996

Degradative pathways for p-toluenecarboxylate and p-toluenesulfonate and their multicomponent oxygenases in Comamonas testosteroni strains PSB-4 and T-2.

Three multicomponent oxygenases involved in the degradation of p-toluenesulfonate and p-toluenecarboxylate and the regulation of their synthesis have been examined in three strains (T-2, PSB-4 and TER-1) of Comamonas testosteroni. Strain T-2 utilizes p-toluenesulfonate as a source of carbon and energy for growth via p-sulfobenzoate and protocatechuate, and p-toluenecarboxylate via terephthalate and protocatechuate, and has the unusual property of requiring the reductase (TsaB) of the toluenesulfonate methyl monooxygenase system (TsaMB) in an incompletely expressed sulfobenzoate dioxygenase system (PsbAC) [Schläfli Oppenberg, H.R., Chen, G., Leisinger, T. & Cook, A. M. (1995). Microbiology 141, 1891-1899]. The independently isolated C. testosteroni PSB-4 utilized only sulfobenzoate and terephthalate via protocatechuate. Mutant TER-1, derived from strain T-2, utilized only terephthalate via protocatechuate. We detected no enzymes of the pathway from toluenesulfonate to sulfobenzoate in strains PSB-4 and TER-1, and confirmed by PCR and Southern blot analysis that the genes (tsaMB) encoding toluenesulfonate monooxygenase were absent. We concluded that, in strain PSB-4, the regulatory unit encoding the genes for the conversion of toluenesulfonate to sulfobenzoate was missing, and that generation of mutant TER-1 involved deletion of this regulatory unit and of the regulatory unit encoding desulfonation of sulfobenzoate. The degradation of sulfobenzoate in strain PSB-4 was catalysed by a fully inducible sulfobenzoate dioxygenase system (PsbACPSB-4), which, after purification of the oxygenase component (PsbAPSB-4), turned out to be indistinguishable from the corresponding component from strain T-2 (PsbAT-2). Reductase PsbCPSB-4, which we could separate but not purify, was active with oxygenase PsbAPSB-4 and PsbAT-2. Oxygenase PsbAPSB-4 was shown by electron paramagnetic resonance spectroscopy to contain a Rieske [2Fe-2S] centre. The enzyme system oxygenating terephthalate was examined and the oxygenase component purified and characterized. The oxygenase component in strains T-2 (and mutant TER-1) and PSB-4 were indistinguishable. The reductase component, which we separated but failed to purify, was active with the oxygenase from all strains. Gains and losses of blocks of genes in evolution is discussed.

Benzenesulfonates↗

PubMed1996

The assimilation of sulfur from multiple sources and its correlation with expression of the sulfate-starvation-induced stimulon in Pseudomonas putida S-313.

Conditions were optimized for the batch growth of Pseudomonas putida S-313 under sulfur-limited conditions. P. putida grew exponentially with sulfate as the sole source of sulfur, and growth was concomitant with the utilization of sulfate until it was exhausted. A further 20% of protein was synthesized after the apparent disappearance of sulfate. A mass balance for the utilized sulfate in cell material was calculated, given the observed molar growth yield of about 3.6 kg protein (mol S)-1 and a sulfur content of 0.41% S in dry matter. Similar data were obtained for growth with cysteine and thiocyanate. The organism also grew exponentially with 4-toluenesulfonate (TS) as sulfur source, essentially as observed with sulfate, except that negligible protein formation after exhaustion of TS was observed. Similar data were also obtained with 4-nitrocatecholsulfate (NCS) and ethanesulfonate. Any substrate pair selected from sulfate, cysteine and thiocyanate was utilized simultaneously, and although one of the pair of substrates was always preferred, growth continued at the same rate when only one substrate remained. Growth after substrate exhaustion was observed. Any substrate pair selected from TS, NCS and ethanesulfonate gave similar data, but with less growth after exhaustion of the sulfur sources. If a mixed substrate pair was chosen from the two groups, the sulfur source from the first-named group was initially used exclusively, and the second source of sulfur was utilized subsequently, after a lag phase. The data are considered to reflect the control of scavenging for sulfur and of distribution of sulfur in the cell exerted by the sulfate-starvation-induced stimulon.

Culture Media↗

PubMed1996

Identification of sulfate starvation-regulated genes in Escherichia coli: a gene cluster involved in the utilization of taurine as a sulfur source.

Genes whose expression is regulated by sulfate starvation in Escherichia coli were identified by generating random translational lacZ fusions in the chromosome with the lambda placMu9 system. Nine lacZ fusion strains which expressed beta-galactosidase after growth under sulfate starvation conditions but not after growth in the presence of sulfate were found. These included two strains with insertions in the dmsA and rhsD genes, respectively, and seven strains in which the insertions were located within a 1.8-kb region downstream of hemB at 8.5 minutes on the E. coli chromosome. Analysis of the nucleotide sequence of this region indicated the presence of four open reading frames designated tauABCD. Disruption of these genes resulted in the loss of the ability to utilize taurine (2-aminoethanesulfonate) as a source of sulfur but did not affect the utilization of a range of other aliphatic sulfonates as sulfur sources. The TauA protein contained a putative signal peptide for transport into the periplasm; the TauB and TauC proteins showed sequence similarity to ATP-binding proteins and membrane proteins, respectively, of ABC-type transport systems; and the TauD protein was related in sequence to a dichlorophenoxyacetic acid dioxygenase. We therefore suggest that the proteins encoded by tauABC constitute an uptake system for taurine and that the product of tauD is involved in the oxygenolytic release of sulfite from taurine. The transcription initiation site was detected 26 to 27 bp upstream of the translational start site of tauA. Expression of the tauD gene was dependent on CysB, the transcriptional activator of the cysteine regulon.

Amino Acid Sequence↗

PubMed1996

Coenzyme F390 synthetase from Methanobacterium thermoautotrophicum Marburg belongs to the superfamily of adenylate-forming enzymes.

Depending on the reduction-oxidation state of the cell, some methanogenic bacteria synthesize or hydrolyze 8-hydroxyadenylylated coenzyme F420 (coenzyme F390). These two reactions are catalyzed by coenzyme F390 synthetase and hydrolase, respectively. To gain more insight into the mechanism of the former reaction, coenzyme F390 synthetase from Methanobacterium thermoautotrophicum Marburg was purified 89-fold from cell extract to a specific activity of 0.75 mumol.min-1.mg of protein-1. The monomeric enzyme consisted of a polypeptide with an apparent molecular mass of 41 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. ftsA, the gene encoding coenzyme F390 synthetase, was cloned and sequenced. It encoded a protein of 377 amino acids with a predicted M(r) of 43,280. FtsA was found to be similar to domains found in the superfamily of peptide synthetases and adenylate-forming enzymes. FtsA was most similar to gramicidin S synthetase II (67% similarity in a 227-amino-acid region) and sigma-(L-alpha-aminoadipyl)-L-cysteine-D-valine synthetase (57% similarity in a 193-amino-acid region). Coenzyme F390 synthetase, however, holds an exceptional position in the superfamily of adenylate-forming enzymes in that it does not activate a carboxyl group of an amino or hydroxy acid but an aromatic hydroxyl group of coenzyme F420.

Amino Acid Sequence↗

PubMed1995

Evidence for a defective prophage on the chromosome of Methanobacterium wolfei.

Evidence shows the presence on the chromosome of Methanobacterium wolfei of a defective prophage which, by DNA-DNA hybridization, is closely related to the virulent archaeophage psi M1 of Methanobacterium thermoautotrophicum Marburg. Partial sequencing of a M. wolfei 16S rRNA gene and phylogenetic analysis indicated that this organism is more closely related to other representatives of the genus Methanobacterium than to M. thermoautotrophicum Marburg. The chromosomal region of M. wolfei encoding the putative prophage was found to be deleted for two non-contiguous segments of the phage psi M1 genome and thus encompassed only 80 to 90% of the psi M1 DNA. The prophage region was mapped to a 30 kb restriction fragment on the physical map of the M. wolfei chromosome. A randomly chosen DNA fragment was cloned from phage psi M1 DNA, as was its homologous counterpart from the chromosome of M. wolfei. The 126-bp region present in both clones exhibited 100% sequence identity.

Bacteriophages↗