Limits...
Multiple amino acid sequence alignment nitrogenase component 1: insights into phylogenetics and structure-function relationships.

Howard JB, Kechris KJ, Rees DC, Glazer AN - PLoS ONE (2013)

Bottom Line: Three species in Group Nif-III, Candidatus Desulforudis audaxviator, Desulfotomaculum kuznetsovii, and Thermodesulfatator indicus, were found to have a seleno-cysteine that replaces one cysteinyl ligand of the 8Fe:7S, P-cluster.Subsets of invariant residues, limited to individual groups, were identified; these unique residues help identify the gene of origin (anf, nif, or vnf) yet should not be considered diagnostic of the metal content of associated cofactors.Fourteen of the 19 residues that compose the cofactor pocket are invariant or single variant; the other five residues are highly variable but do not correlate with the putative metal content of the cofactor.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minnesota, United States of America ; Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California, United States of America.

ABSTRACT
Amino acid residues critical for a protein's structure-function are retained by natural selection and these residues are identified by the level of variance in co-aligned homologous protein sequences. The relevant residues in the nitrogen fixation Component 1 α- and β-subunits were identified by the alignment of 95 protein sequences. Proteins were included from species encompassing multiple microbial phyla and diverse ecological niches as well as the nitrogen fixation genotypes, anf, nif, and vnf, which encode proteins associated with cofactors differing at one metal site. After adjusting for differences in sequence length, insertions, and deletions, the remaining >85% of the sequence co-aligned the subunits from the three genotypes. Six Groups, designated Anf, Vnf , and Nif I-IV, were assigned based upon genetic origin, sequence adjustments, and conserved residues. Both subunits subdivided into the same groups. Invariant and single variant residues were identified and were defined as "core" for nitrogenase function. Three species in Group Nif-III, Candidatus Desulforudis audaxviator, Desulfotomaculum kuznetsovii, and Thermodesulfatator indicus, were found to have a seleno-cysteine that replaces one cysteinyl ligand of the 8Fe:7S, P-cluster. Subsets of invariant residues, limited to individual groups, were identified; these unique residues help identify the gene of origin (anf, nif, or vnf) yet should not be considered diagnostic of the metal content of associated cofactors. Fourteen of the 19 residues that compose the cofactor pocket are invariant or single variant; the other five residues are highly variable but do not correlate with the putative metal content of the cofactor. The variable residues are clustered on one side of the cofactor, away from other functional centers in the three dimensional structure. Many of the invariant and single variant residues were not previously recognized as potentially critical and their identification provides the bases for new analyses of the three-dimensional structure and for mutagenesis studies.

Show MeSH
Phylogeny of species used for multi-sequence alignment of NifD and NifK.The species in the data analysis set (identifiers and species are in Table S1) were superimposed on a simplified whole-proteome tree from Jun et al. (Figure 2 in [34], constructed with whole proteomes of 884 prokaryotes). Identifiers are based upon the six nitrogenase groups; species with both Nif and either Anf or Vnf have more than one identifier.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3760896&req=5

pone-0072751-g002: Phylogeny of species used for multi-sequence alignment of NifD and NifK.The species in the data analysis set (identifiers and species are in Table S1) were superimposed on a simplified whole-proteome tree from Jun et al. (Figure 2 in [34], constructed with whole proteomes of 884 prokaryotes). Identifiers are based upon the six nitrogenase groups; species with both Nif and either Anf or Vnf have more than one identifier.

Mentions: For critical residues to be revealed by natural selection, a fundamental requirement is that the species used in the multiple sequence alignment represent a broad, distinctive phylogenetic distribution. Although the number of known species with putative nitrogen fixation genes greatly exceeds the 75 species used here (e.g., [33]), the criteria for inclusion of the species were that whole genomes are available, that a broad range of classes is represented, and that the species exemplify metabolic diversity and distinctive ecological niches. One goal of this study is to correlate the sequences of the three known genetic variants of nitrogenase which also have different apparent metal requirements in the cofactor. When Anf and Vnf versions of Component 1 were available, the Nif sequences from the same species were included. The diversity of species in our analysis is indicated by the distribution of these species across nearly the whole proteome map of Jun et al. [34] as shown in Figure 2. Their tree was constructed based on analyzing 884 full genomes and independent of the ability of a species to fix nitrogen. For our purpose, we have superimposed the species from our study on a simplified version of their map to show the distribution in the larger microbial world. A second demonstration of the species distribution is shown in Figure S1 constructed independently using the 16S rRNA similarity index for just the species in our data set. Jun et al. [34] observed that, with some important exceptions, there is good agreement between these two types of maps of the microbial world. However, we found some potentially interesting differences when the nitrogen fixation genes are considered. These differences may reflect the lower resolution of the 16S rRNA map as well as horizontal gene transfer [4].


Multiple amino acid sequence alignment nitrogenase component 1: insights into phylogenetics and structure-function relationships.

Howard JB, Kechris KJ, Rees DC, Glazer AN - PLoS ONE (2013)

Phylogeny of species used for multi-sequence alignment of NifD and NifK.The species in the data analysis set (identifiers and species are in Table S1) were superimposed on a simplified whole-proteome tree from Jun et al. (Figure 2 in [34], constructed with whole proteomes of 884 prokaryotes). Identifiers are based upon the six nitrogenase groups; species with both Nif and either Anf or Vnf have more than one identifier.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3760896&req=5

pone-0072751-g002: Phylogeny of species used for multi-sequence alignment of NifD and NifK.The species in the data analysis set (identifiers and species are in Table S1) were superimposed on a simplified whole-proteome tree from Jun et al. (Figure 2 in [34], constructed with whole proteomes of 884 prokaryotes). Identifiers are based upon the six nitrogenase groups; species with both Nif and either Anf or Vnf have more than one identifier.
Mentions: For critical residues to be revealed by natural selection, a fundamental requirement is that the species used in the multiple sequence alignment represent a broad, distinctive phylogenetic distribution. Although the number of known species with putative nitrogen fixation genes greatly exceeds the 75 species used here (e.g., [33]), the criteria for inclusion of the species were that whole genomes are available, that a broad range of classes is represented, and that the species exemplify metabolic diversity and distinctive ecological niches. One goal of this study is to correlate the sequences of the three known genetic variants of nitrogenase which also have different apparent metal requirements in the cofactor. When Anf and Vnf versions of Component 1 were available, the Nif sequences from the same species were included. The diversity of species in our analysis is indicated by the distribution of these species across nearly the whole proteome map of Jun et al. [34] as shown in Figure 2. Their tree was constructed based on analyzing 884 full genomes and independent of the ability of a species to fix nitrogen. For our purpose, we have superimposed the species from our study on a simplified version of their map to show the distribution in the larger microbial world. A second demonstration of the species distribution is shown in Figure S1 constructed independently using the 16S rRNA similarity index for just the species in our data set. Jun et al. [34] observed that, with some important exceptions, there is good agreement between these two types of maps of the microbial world. However, we found some potentially interesting differences when the nitrogen fixation genes are considered. These differences may reflect the lower resolution of the 16S rRNA map as well as horizontal gene transfer [4].

Bottom Line: Three species in Group Nif-III, Candidatus Desulforudis audaxviator, Desulfotomaculum kuznetsovii, and Thermodesulfatator indicus, were found to have a seleno-cysteine that replaces one cysteinyl ligand of the 8Fe:7S, P-cluster.Subsets of invariant residues, limited to individual groups, were identified; these unique residues help identify the gene of origin (anf, nif, or vnf) yet should not be considered diagnostic of the metal content of associated cofactors.Fourteen of the 19 residues that compose the cofactor pocket are invariant or single variant; the other five residues are highly variable but do not correlate with the putative metal content of the cofactor.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minnesota, United States of America ; Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California, United States of America.

ABSTRACT
Amino acid residues critical for a protein's structure-function are retained by natural selection and these residues are identified by the level of variance in co-aligned homologous protein sequences. The relevant residues in the nitrogen fixation Component 1 α- and β-subunits were identified by the alignment of 95 protein sequences. Proteins were included from species encompassing multiple microbial phyla and diverse ecological niches as well as the nitrogen fixation genotypes, anf, nif, and vnf, which encode proteins associated with cofactors differing at one metal site. After adjusting for differences in sequence length, insertions, and deletions, the remaining >85% of the sequence co-aligned the subunits from the three genotypes. Six Groups, designated Anf, Vnf , and Nif I-IV, were assigned based upon genetic origin, sequence adjustments, and conserved residues. Both subunits subdivided into the same groups. Invariant and single variant residues were identified and were defined as "core" for nitrogenase function. Three species in Group Nif-III, Candidatus Desulforudis audaxviator, Desulfotomaculum kuznetsovii, and Thermodesulfatator indicus, were found to have a seleno-cysteine that replaces one cysteinyl ligand of the 8Fe:7S, P-cluster. Subsets of invariant residues, limited to individual groups, were identified; these unique residues help identify the gene of origin (anf, nif, or vnf) yet should not be considered diagnostic of the metal content of associated cofactors. Fourteen of the 19 residues that compose the cofactor pocket are invariant or single variant; the other five residues are highly variable but do not correlate with the putative metal content of the cofactor. The variable residues are clustered on one side of the cofactor, away from other functional centers in the three dimensional structure. Many of the invariant and single variant residues were not previously recognized as potentially critical and their identification provides the bases for new analyses of the three-dimensional structure and for mutagenesis studies.

Show MeSH