Substrate Specificity and Kinetic Studies of Nodulation Protein NodL of Rhizobium leguminosarum

Gv, Bloemberg; Rm, Lagas; S, van Leeuwen; Ga, van der Marel; Jh, van Boom; Bj, Lugtenberg; Hp, Spaink

doi:10.1021/bi00039a030

Cited by 28 publications

(16 citation statements)

References 19 publications

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“…NodA, NodB, and NodC proteins are required for the synthesis of the acylated Nod factor backbone, whereas most other Nod proteins are required for specific modifications (6,7). NodL of Rhizobium leguminosarum is an acetyltransferase (8), and nodZ encodes an ␣-1,6-fucosyltransferase (9 -11). Sulfation of Nod factors produced by Sinorhizobium meliloti is carried out by the sulfotransferase NodH that uses 3Ј-phosphoadenosine 5Ј-phosphosulfate, synthesized by the NodPQ complex (12,13).…”

mentioning

confidence: 99%

Nod Factor Requirements for Efficient Stem and Root Nodulation of the Tropical Legume Sesbania rostrata

D’Haeze¹,

Mergaert²,

Promé³

et al. 2000

Journal of Biological Chemistry

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Azorhizobium caulinodans ORS571 synthesizes mainly pentameric Nod factors with a household fatty acid, an N-methyl, and a 6-O-carbamoyl group at the nonreducing-terminal residue and with a D-arabinosyl, an L-fucosyl group, or both at the reducing-terminal residue. Nodulation on Sesbania rostrata was carried out with a set of bacterial mutants that produce well characterized Nod factor populations. Purified Nod factors were tested for their capacity to induce root hair formation and for their stability in an in vitro degradation assay with extracts of uninfected adventitious rootlets. The glycosylations increased synergistically the nodulation efficiency and the capacity to induce root hairs, and they protected the Nod factor against degradation. The D-arabinosyl group was more important than the L-fucosyl group for nodulation efficiency. Replacement of the 6-O-L-fucosyl group by a 6-O-sulfate ester did not affect Nod factor stability, but reduced nodulation efficiency, indicating that the L-fucosyl group may play a role in recognition. The 6-O-carbamoyl group contributes to nodulation efficiency, biological activity, and protection, but could be replaced by a 6-O-acetyl group for root nodulation. The results demonstrate that none of the studied substitutions is strictly required for triggering normal nodule formation. However, the nodulation efficiency was greatly determined by the synergistic presence of substitutions. Within the range tested, fluctuations of Nod factor amounts had little impact on the symbiotic phenotype.

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mentioning

confidence: 99%

Nod Factor Requirements for Efficient Stem and Root Nodulation of the Tropical Legume Sesbania rostrata

D’Haeze¹,

Mergaert²,

Promé³

et al. 2000

Journal of Biological Chemistry

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“…It is known that NodL acts preferentially on the NodBC metabolite rather than on a more mature acyl-bearing species (2). From our observations, we suggest that NodS-mediated N methylation of such NodBCL metabolites may well be impossible and that N-methylated, nonreducing-terminally O-acety-lated LCOs would therefore be very unlikely to be biosynthesized.…”

Section: Discussionmentioning

confidence: 76%

“…Fully N-acetylated chitooligosaccharides or unmethylated LCOs are not methylated in vitro by NodS (11). In constrast, the NodL protein is able to acetylate, in addition to terminally de-N-acetylated chitooligosaccharides, various other substrates, such as LCOs, chitin oligosaccharides, chitosan oligosaccharides, N-acetylglucosamine, and cellopentaose (2,3). From all these data it is clear that NodS is highly substrate specific while NodL has a broad substrate specificity.…”

Section: Discussionmentioning

confidence: 96%

“…NodS uses N-deacetylated chitooligosaccharides, the products of the NodBC proteins (the so-called NodBC metabolites), as its methyl acceptors (11,18). Although the NodL protein appears to be able to acetylate various substrates, such as LCOs, chitin fragments, and N-acetylglucosamine, the NodBC metabolite is the preferred substrate and is probably the in vivo acetyl-accepting substrate (2,3).…”

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confidence: 99%

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Rhizobial NodL O -Acetyl Transferase and NodS N -Methyl Transferase Functionally Interfere in Production of Modified Nod Factors

et al. 2001

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The products of the rhizobial nodulation genes are involved in the biosynthesis of lipochitin oligosaccharides (LCOs), which are host-specific signal molecules required for nodule formation. The presence of an O-acetyl group on C-6 of the nonreducing N-acetylglucosamine residue of LCOs is due to the enzymatic activity of NodL. Here we show that transfer of the nodL gene into four rhizobial species that all normally produce LCOs that are not modified on C-6 of the nonreducing terminal residue results in production of LCOs, the majority of which have an acetyl residue substituted on C-6. Surprisingly, in transconjugant strains of Mesorhizobium loti, Rhizobium etli, and Rhizobium tropici carrying nodL, such acetylation of LCOs prevents the endogenous nodS-dependent transfer of the N-methyl group that is found as a substituent of the acylated nitrogen atom. To study this interference between nodL and nodS, we have cloned the nodS gene of M. loti and used its product in in vitro experiments in combination with purified NodL protein. It has previously been shown that a chitooligosaccharide N deacetylated on the nonreducing terminus (the so-called NodBC metabolite) is the preferred substrate for NodS as well as for NodL. Here we show that the NodBC metabolite, acetylated by NodL, is not used by the NodS protein as a substrate while the NodL protein can acetylate the NodBC metabolite that has been methylated by NodS.Rhizobial bacteria have the unique ability to induce formation of nitrogen-fixing nodules on the roots or stems of leguminous plants. The development of legume nodules is largely controlled by reciprocal signal exchange between the symbiotic partners. Legume roots secrete specific flavonoids or isoflavonoids that induce the transcription of many bacterial genes (nod, nol, and noe genes). Most of these genes are involved in the synthesis and secretion of signal molecules that are essential to trigger nodule formation and that are known as Nod factors. Nod factors from many rhizobial species have been characterized, and their structures have been elucidated. Because their basic structure consists of a chitin oligosaccharide backbone N acylated on the nonreducing terminal residue, they are referred to as lipochitin oligosaccharides (LCOs). The nature of the fatty acid and the combination of diverse chemical substitutions provide host specificity to the LCOs produced by a given rhizobial strain (for reviews, see references 1, 7, 9, and 25).

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“…1). A homology search for the pssR gene product revealed similarity of its C-terminal region (amino acids 87-136) with the corresponding parts of a large number of acetyltransferases, including well characterized rhizobial NodL [71][72][73] Notably pssR orthologs were found in all PssI-clusters (Fig. 1).…”

Section: Genes Involved In Modification Of Epsmentioning

confidence: 99%

Exopolysaccharide Biosynthesis in Rhizobium leguminosarum: From Genes to Functions

Ивашина¹,

Ksenzenko²

2012

The Complex World of Polysaccharides

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Substrate Specificity and Kinetic Studies of Nodulation Protein NodL of Rhizobium leguminosarum

Cited by 28 publications

References 19 publications

Nod Factor Requirements for Efficient Stem and Root Nodulation of the Tropical Legume Sesbania rostrata

Nod Factor Requirements for Efficient Stem and Root Nodulation of the Tropical Legume Sesbania rostrata

Rhizobial NodL O -Acetyl Transferase and NodS N -Methyl Transferase Functionally Interfere in Production of Modified Nod Factors

Exopolysaccharide Biosynthesis in Rhizobium leguminosarum: From Genes to Functions

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