Rhizobium sp. strain ORS571 grows synergistically on N2 and nicotinate as N sources

Ludwig, Robert A.

doi:10.1128/jb.165.1.304-307.1986

Cited by 18 publications

(18 citation statements)

References 11 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, because it mnust be supplemented with nicotinate, which is catabolized under N2-fixing conditions, A. caulinodans is not a diazotroph. Rather, it conducts synergistic N2 fixation and nicotinate catabolism (16), as evidenced by the following experiments. When N2-fixing A. caulinodans cultures under an atmosphere of 97% N2-3% 02 are supplemented with 10-fold-augmented nicotinate levels (16 to 160 ,uM), a 10-fold increase in cell yield occurs.…”

mentioning

confidence: 99%

Identification of cyclic intermediates in Azorhizobium caulinodans nicotinate catabolism

et al. 1989

Self Cite

View full text Add to dashboard Cite

In wild-type Azorhizobium caulinodans ORS571, nicotinate served both as anabolic substrate for NAD+ production and as catabolic substrate for use as the N source. Catabolic enzyme activities were greatest from cultures grown with nicotinate as the N source and least when cultures were grown with ammonium as the N source. Vector insertion mutants unable to catabolize nicotinate (nic::Vi mutants) still required micromolar quantities of this compound for growth. Therefore, A. caulinodans wild type is NAD+ auxotrophic. As the first two intermediates in A. caulinodans nicotinate catabolism, two cyclic compounds, 6-hydroxynicotinate and 1,4,5,6-tetrahydro-6-oxonicotinate, were identified. These compounds were purified from the growth medium of strain 61009 (a nic::Vi mutant) by high-performance liquid chromatography; their identities were subsequently confirmed by UV absorbance, nuclear magnetic resonance, and mass spectra. The conversion of 1 mol of nicotinate to 6-hydroxynicotinate consumed 0.5 mol of 02. From 180 isotopic incorporation experiments, water was the hydroxyl-equivalent source. A nicotinate hydroxylase activity proved to be cell wall-membrane associated; this activity served as direct electron donor (not indirect via NADP+) to 02 via membrane electron transport. These catabolic reactions have not previously been witnessed together in the same organism. A. caulinodans nicotinate catabolism seems coupled to N2 fixation, although the explicit mechanism of this coupling remains to be determined.

show abstract

mentioning

confidence: 99%

Identification of cyclic intermediates in Azorhizobium caulinodans nicotinate catabolism

et al. 1989

Self Cite

View full text Add to dashboard Cite

show abstract

“…Although not demonstrated, Azorhizobium GCADH activity presumably also yields crotonyl-CoA (Fig. 1) Because it requires nicotinate for NAD+ synthesis, and because under optimum N2-fixing physiological conditions it rapidly catabolizes nicotinate as an N source, the wild-type Azorhizobium strain is not a true diazotroph (22). In Azorhizobium N2-fixing chemostat cultures, less than 10% of the assimilated N culture yield was estimated as derived from nicotinate N; more than 90% was derived from fixed N2 (3,12).…”

Section: Discussionmentioning

confidence: 99%

“…All wild Azorhizobium isolates are phenotypically indistinguishable and are auxotrophic for pyridine nucleotide cofactor (NAD+) biosynthesis; culture in defined medium requires nicotinate supplementation (10). When so cultured, A. caulinodans also uses nicotinate as the sole N source for growth (22). Whereas all other studied rhizobia exclusively fix N2 in symbiosis, A. caulinodans avidly fixes N2 in culture (10).…”

mentioning

confidence: 99%

Elucidation of the complete Azorhizobium nicotinate catabolism pathway

et al. 1992

Self Cite

View full text Add to dashboard Cite

A complete pathway for Azorhizobium caulinodans nicotinate catabolism has been determined from mutant phenotype analyses, isolation of metabolic intermediates, and structural studies. Nicotinate serves as a respiratory electron donor to 02 via a membrane-bound hydroxylase and a specific c-type cytochrome oxidase. The resulting oxidized product, 6-hydroxynicotinate, is next reduced to 1,4,5,6-tetrahydro-6-oxonicotinate. Hydrolytic ring breakage follows, with release of pyridine N as ammonium. Decarboxylation then releases the nicotinate C-7 carboxyl group as C02, and the remaining C skeleton is then oxidized to yield glutarate. Transthioesterification with succinyl coenzyme A (succinyl-CoA) yields glutaryl-CoA, which is then oxidatively decarboxylated to yield crotonyl-CoA. As with general acyl x oxidation, L-P-hydroxybutyryl-CoA, acetoacetylCoA, and finally two molecules of acetyl-CoA are produced. In sum, nicotinate is catabolized to yield two CO2 molecules, two acetyl-CoA molecules, and ammonium. Nicotinate catabolism stimulates Azorhizobium N2 fixation rates in culture. Nicotinate catabolism mutants still able to liberate pyridine N as ammonium retain this capability, whereas mutants so blocked do not. From, mutant analyses and additional physiological tests, N2 fixation stimulation is indirect. In N-limited culture, nicotinate catabolism augments anabolic N pools and, as a consequence, yields N2-fixing cells with higher dinitrogenase content.Azorhizobium caulinodans invades both stems and roots of the host legume Sesbania rostrata and yields symbiotic nodules that fix N2 at very high rates. All wild Azorhizobium isolates are phenotypically indistinguishable and are auxotrophic for pyridine nucleotide cofactor (NAD+) biosynthesis; culture in defined medium requires nicotinate supplementation (10). When so cultured, A. caulinodans also uses nicotinate as the sole N source for growth (22). Whereas all other studied rhizobia exclusively fix N2 in symbiosis, A. caulinodans avidly fixes N2 in culture (10). Paradoxically, when fixing N2 in culture under optimal conditions, A. caulinodans exhaustively catabolizes nicotinate, and its growth becomes NAD+ limited. Relative to a submicromolar nicotinate vitamin requirement, high-level (0.3 mM) nicotinate supplementation strongly stimulates N2 fixation in culture (4, 11). We have sought to understand how nicotinate stimulates N2 fixation.Although nicotinate is a good N source, it is a poor C source for Azorhizobium growth (18). However, stoichiometric measurements of nicotinate-stimulated 02 consumption indicate that A. caulinodans completely utilizes the nicotinate C skeleton (18). Cyclic intermediates in this pathway were previously isolated and characterized, and a mechanism of pyridine ring N release was proposed (18) (Fig. 1). Nicotinate is first oxidized to yield 6-hydroxynicotinate by a membrane-bound hydroxylase, which uses 02 as an electron acceptor. The nicotinate-specific respiratory chain to 02 terminates at a c-type cytochrome oxidase (17). The reaction prod...

show abstract

“…Subsequently, the complex growth interdependence of N 2 -fixing A. caulinodans cultures and nicotinate was investigated in depth (3, 10-12). Importantly, A. caulinodans fails to synthesize pyridine nucleotide cofactors de novo and thus requires nicotinate (vitamin) supplementation; A. caulinodans is thus auxotrophic for NAD ϩ biosynthesis (11).Apart from nicotinate's fate as an anabolic substrate, nicotinate catabolism seems to somehow stimulate N 2 fixation in culture (3,(10)(11)(12). Nicotinate is first oxidized to 6-OH-nicotinate by a membrane-bound nicotinate hydroxylase (11).…”

mentioning

confidence: 99%

“…Apart from nicotinate's fate as an anabolic substrate, nicotinate catabolism seems to somehow stimulate N 2 fixation in culture (3,(10)(11)(12). Nicotinate is first oxidized to 6-OH-nicotinate by a membrane-bound nicotinate hydroxylase (11).…”

mentioning

confidence: 99%

Nicotinate catabolism is dispensable and nicotinate anabolism is crucial in Azorhizobium caulinodans growing in batch culture and chemostat culture on N2 as The N source

et al. 1995

View full text Add to dashboard Cite

When Azorhizobium caulinodans was grown in chemostat cultures with N 2 as the N source at a constant dilution rate of 0.1 h ؊1 in media with a constant concentration (50 mM) of succinate and variable concentrations (1.5 to 585 M) of nicotinate, neither the growth yield on succinate, the specific rate of O 2 consumption, nor the specific rate of CO 2 production showed linear regression with the concentration of nicotinate. Azorhizobium caulinodans is unique among rhizobia because not only is it able to reduce N 2 to NH 3 , but it is also able to incorporate the NH 3 so formed into biomass in pure culture. The assimilation of NH 3 occurs very efficiently. Under several physiological conditions conducive to NH 3 excretion, no NH 3 diffused out of bacterial cells (8). To date, among the rhizobia, real growth on N 2 in free-living cultures has been observed only for A. caulinodans. In this respect, it is intermediate between the symbiotic and free-living N 2 -fixing bacteria.The prerequisites for N 2 fixation are a low dissolved O 2 tension (DOT) and the absence of combined nitrogen. For growth of A. caulinodans in defined medium, the vitamins biotin, pantothenate, and nicotinate are essential (5). When these vitamin requirements were reexamined by measuring the maximal amount of biomass (by the absorbance at 570 nm [A 570 ]) produced in batch cultures, only biotin and pantothenate were needed in (sub)micromolar quantities, whereas nicotinate was required at relatively high (0.3 mM) levels (7). Subsequently, the complex growth interdependence of N 2 -fixing A. caulinodans cultures and nicotinate was investigated in depth (3, 10-12). Importantly, A. caulinodans fails to synthesize pyridine nucleotide cofactors de novo and thus requires nicotinate (vitamin) supplementation; A. caulinodans is thus auxotrophic for NAD ϩ biosynthesis (11).Apart from nicotinate's fate as an anabolic substrate, nicotinate catabolism seems to somehow stimulate N 2 fixation in culture (3,(10)(11)(12). Nicotinate is first oxidized to 6-OH-nicotinate by a membrane-bound nicotinate hydroxylase (11). When supplemented with 0.3 mM 6-OH-nicotinate, mutants defective in nicotinate hydroxylase fixed N 2 at the wild-type level (3). Recently, the complete A. caulinodans nicotinate catabolism pathway has been elucidated by analysis of mutants affected in various steps of this pathway (10). Nicotinate catabolism mutants still able to liberate pyridine N as ammonium retained the capability to enhance acetylene reduction rates in nitrogenase induction medium by increased nicotinate supplementation. It was concluded that nicotinate catabolism indirectly stimulated N 2 fixation by augmenting anabolic N pools in Nlimited cultures (10), but at the same time neither nicotinate nor 6-OH-nicotinate catabolism seemed to be essential for N 2 fixation (3, 11).The above studies were all carried out with poorly defined batch cultures or in nitrogenase induction media in which only the onset of acetylene reduction was measured. Vitamin requirements of A. caulinodans co...

show abstract

Rhizobium sp. strain ORS571 grows synergistically on N2 and nicotinate as N sources

Cited by 18 publications

References 11 publications

Identification of cyclic intermediates in Azorhizobium caulinodans nicotinate catabolism

Identification of cyclic intermediates in Azorhizobium caulinodans nicotinate catabolism

Elucidation of the complete Azorhizobium nicotinate catabolism pathway

Nicotinate catabolism is dispensable and nicotinate anabolism is crucial in Azorhizobium caulinodans growing in batch culture and chemostat culture on N2 as The N source

Contact Info

Product

Resources

About