Reductive cleavage of sarcosine and betaine by Eubacterium acidaminophilum via enzyme systems different from glycine reductase

Hormann, K.; Andreesen, Jan R.

doi:10.1007/bf00277541

Cited by 81 publications

(62 citation statements)

References 43 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Therefore, the formate dehydrogenase was also purified by a similar procedure from crude extract of sarcosine-grown cells. However, the formate dehydrogenase activity-containing fraction finally obtained had no P, or P, sarCO,,,,e activity in the respective complementation assay and no bands at 45, 48, and 83 kDa, but other protein bands known from purifications of P, (54 kDa and 44 kDa) [8] and P, (27 kDa and 47 kDa, see below) and a 62-kDa protein were present (Fig. 4A, lane 2).…”

Section: Resultsmentioning

confidence: 95%

Purification and Characterization of Protein P_B of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum

Meyer

Granderath

Andreesen

1995

European Journal of Biochemistry

View full text Add to dashboard Cite

Simple complementation assay systems were developed for the substrate-specific proteins P, of glycine reductase, sarcosine reductase, and betaine reductase, in which acetyl phosphate was detected as the product in all three cases. The betaine-specific subunits of protein B (P, responsible for betaine reductase activity were purified to homogeneity from cells of Eubucterium ueidam~noph~lum. The molecular masses of the two different subunits were 45 kDa and 48 kDa according to SDS/PAGE. The molecular mass of the native protein was about 200 kDa, indicating an u2P2 structure. The glycine-specific protein B (P, gl,c,,J was partially purified and subunits of 47 kDa and 27 kDa were N-terminally sequenced. The latter subunits cross-reacted with antibodies raised against P, and showed high sequence similarity to the 45-kDa and 48-kDa subunits of P, hcf.,inc, respectively. 12-"C]Glycine could be covalently coupled to the 47-kDa subunit by treatment with borohydride. By the same procedure, [2-"C]sarcosine labeled a protein of the same size. Like the sarcosine reductase activity, this protein was not present in glycinegrown cells, indicating its specific involvement in sarcosine metabolism. The labile viologen-dependent formate dehydrogenase purified with the respective P,, proteins and could be tentatively assigned to a 95-kDa protein.Keywords: Eubacterium trcidaminophiluin ; glycine reductase ; sarcosine reductase ; betaine reductase ; formate dehydrogenase.Glycine can be utilized by Eubucterium acidaminophilum as sole carbon source by the action of glycine decarboxylase and glycine reductase 11 1. The oxidation of glycine to S,lO-methylenetetrahydrofolate, ammonia, and reduced pyridine nucleotides is performed by glycine decarboxylase. The dihydrolipoamide dehydrogenase (P,) usually involved in the total reaction is not part of the latter enzyme complex in E. acidurninophilum 12. 31. Its function is taken over by a thioredoxin system, catalyzing the same, but NADP(H)-specific reaction that channels the electrons directly to the glycine reductase proteins 13, 41. The glycine reductase of E. acidurnirzophilum consists of at least three proteins (Scheme 1) : the selenocysteine-containing protein A (PA) ; the substrate-binding protein B (P,) ; the acetyl phosphate-forming protein C (P,) ( 5 -7 1. The 5,1O-methylenetetrahydrofolate formed by glycine oxidation is further oxidized in two steps by methylenetetrahydrofolate dehydrogenase and formate dehydrogenase to CO:. Both enzymes are specific for NADP(H) in E. reductase is also present in cells grown on sarcosine or betaine 181 in contrast to the situation in betaine-grown cells of Closrridium litorule, as revealed in this study, where only a betaine reductase, but no glycine reductase activity could be determined. Antibodies directed against P,, and P,. of E. uciduminophilum grown on glycine cross-react with homologous proteins of E. ncidaminophilurn and C. litorale grown on betaine [S, 61, indi-cating that betaine reduction occurs by an enzyme system analogous to glycine ...

show abstract

Section: Resultsmentioning

confidence: 95%

Purification and Characterization of Protein P_B of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum

Meyer

Granderath

Andreesen

1995

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…During autoclaving, 75% of the creatine may become converted to creatinine (Moller et al, 1986). One millilitre trace element solution SL-A, 1 ml resazurin solution (1 g 1-' ) and 5 ml vitamin stock solution were added per litre of medium as described previously (Hormann & Andreesen, 1989). The following sterilized components were added separately to the autoclaved medium to give the indicated final concentrations : 50 mM NaHCO, and 2.7mM Na,S.…”

Section: Isolation and Cultivation Of Creatinine-degrading Strainsmentioning

confidence: 99%

“…Creatine reductase, sarcosine reductase and glycine reductase were determined by the formation of [14C]acetate from [ 1 -14C]creatine, [2-14C]sarcosine or [2-14C]glycine as described in general by Stadtman (1970) using the modifications of Hormann & Andreesen (1989). In addition, a non-radioactive enzyme assay was used by measuring the formation of acetyl phosphate (Meyer et al, 1995).…”

Section: Isolation and Cultivation Of Creatinine-degrading Strainsmentioning

confidence: 99%

“…Creatine is an important natural substance (Shimizu et al, 1986) which might allow the evolution of a specific reductase of its own, as is found for glycine, sarcosine and betaine (Hormann & Andreesen, 1989;Meyer et al, 1995) but not for dimethylglycine (Andreesen, 1994). For this reason, highly specific conditions were chosen for enrichment with creatinine and formate as substrates.…”

Section: Physiological and Biochemical Aspectsmentioning

confidence: 99%

“…Clostridium paraputrijicum and Clostridium perfringens metabolized creatinine and creatine anaerobically to N-methylhydantoin (Szulmajster, 1958a, b;ten Krooden & Owens, 1957). The creatine-and sarcosine-utilizing Eubacterium acidaminophilum was shown to require selenite and suitable electron donors (Hormann & Andreesen, 1989 ;Zindel et al, 1988). Creatinine and N-methylhydantoin degradation was studied in several species of Clostridium, Eubacterium and Peptostreptococcus (Gauglitz, 1988).…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Tissierella creatinophila sp. nov., a Gram-positive, anaerobic, non-spore-forming, creatinine-fermenting organism

Harms

Schleicher

Collins³

et al. 1998

International Journal of Systematic Bacteriology

Self Cite

View full text Add to dashboard Cite

A strictly anaerobic, Gram-positive, non-spore-forming bacterium was isolated from sewage sludge which grew on creatinine as sole source of carbon and energy. This new isolate, designated strain KRE 4l, totally degraded creatinine via creatine, sarcosine and glycine to the products acetate, monomethylamine, ammonia and carbon dioxide. Growth on creatinine or creatine was seleniumdependent and stimulated by formate, indicating the involvement of a creatine reductase, sarcosine reductase and/or glycine reductase. This was substantiated by the fact that creatine, sarcosine and glycine were reduced by cell-free extracts. Growth on creatinine or creatine was also possible in the absence of formate, but with an increase in doubling time. The new bacterium occurred as rod-shaped cells, which exhibited an angular form (2-6 pm long and 0.7-1.1 pm wide) and showed motility by means of peritrichous flagella. The G+C content of the DNA was 30 mol0/o. Comparative 16s rRNA sequence analysis demonstrated that strain KRE 4T represents a new subline within the genus Tissierella. Due to its very restricted substrate spectrum and the inability of whole cells to utilize sarcosine and glycine as intermediates of creatine breakdown, this organism can be readily separated from currently described species of Tissierella. Therefore, based on the phenotypic and phylogenetic distinctiveness of the new isolate, it is proposed that the bacterium be classified as a new species of the genus Tissierella, Tissierella creatinophila sp. nov. The type strain is KRE 4 (= DSM 69113.

show abstract

Amino acid variants of the HybB membrane subunit of Escherichia coli [NiFe]‐hydrogenase‐2 support a role in proton transfer

2019

View full text Add to dashboard Cite

[NiFe]‐hydrogenase (Hyd) 2 of Escherichia coli has been proposed to generate proton motive force during H2‐oxidation, which it is dependent on if cells are incubated anaerobically with glycerol to drive reverse H2‐production. The integral membrane subunit HybB is required for proton transfer (PT) by Hyd‐2 but has no cofactor. To provide evidence for PT by HybB, we analyzed the roles of conserved amino acid residues in a predicted proton channel. Exchange of conserved residues identified residues Y99, E133, H184, and E228 as mandatory for PT from the cytoplasm and quinol oxidation. In contrast, exchange of W54, D58, or R89 rendered Hyd‐2 uni‐directional and influenced the equilibrium. Our findings show that HybB is the key subunit in PT.

show abstract

Reductive cleavage of sarcosine and betaine by Eubacterium acidaminophilum via enzyme systems different from glycine reductase

Cited by 81 publications

References 43 publications

Purification and Characterization of Protein P_B of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum

Purification and Characterization of Protein P_B of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum

Tissierella creatinophila sp. nov., a Gram-positive, anaerobic, non-spore-forming, creatinine-fermenting organism

Amino acid variants of the HybB membrane subunit of Escherichia coli [NiFe]‐hydrogenase‐2 support a role in proton transfer

Contact Info

Product

Resources

About

Reductive cleavage of sarcosine and betaine by Eubacterium acidaminophilum via enzyme systems different from glycine reductase

Cited by 81 publications

References 43 publications

Purification and Characterization of Protein PB of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum

Purification and Characterization of Protein PB of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum

Tissierella creatinophila sp. nov., a Gram-positive, anaerobic, non-spore-forming, creatinine-fermenting organism

Amino acid variants of the HybB membrane subunit of Escherichia coli [NiFe]‐hydrogenase‐2 support a role in proton transfer

Contact Info

Product

Resources

About

Purification and Characterization of Protein P_B of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum

Purification and Characterization of Protein P_B of Betaine Reductase and its Relationship to the Corresponding Proteins Glycine Reductase and Sarcosine Reductase from Eubacterium Acidaminophilum