Abstract
BackgroundThe protein encoded by the SA1388 gene from Staphylococcus aureus was chosen for structure determination to elucidate its domain organization and confirm our earlier remote homology based prediction that it housed a nitrogen regulatory PII protein-like domain. SA1388 was predicted to contain a central PII-like domain and two flanking regions, which together belong to the NIF3-like protein family. Proteins like SA1388 remain a poorly studied group and their structural characterization could guide future investigations aimed at understanding their function.ResultsThe structure of SA1388 has been solved to 2.0Å resolution by single wavelength anomalous dispersion phasing method using selenium anomalous signals. It reveals a canonical NIF3-like fold containing two domains with a PII-like domain inserted in the middle of the polypeptide. The N and C terminal halves of the NIF3-like domains are involved in dimerization, while the PII domain forms trimeric contacts with symmetry related monomers. Overall, the NIF3-like domains of SA1388 are organized as a hexameric toroid similar to its homologs, E. coli ybgI and the hypothetical protein SP1609 from Streptococcus pneumoniae. The openings on either side of the toroid are partially covered by trimeric "lids" formed by the PII domains. The junction of the two NIF3 domains has two zinc ions bound at what appears to be a histidine rich active site. A well-defined electron density corresponding to an endogenously bound ligand of unknown identity is observed in close proximity to the metal site.ConclusionSA1388 is the third member of the NIF3-like family of proteins to be structurally characterized, the other two also being hypothetical proteins of unknown function. The structure of SA1388 confirms our earlier prediction that the inserted domain that separates the two NIF3 domains adopts a PII-like fold and reveals an overall capped toroidal arrangement for the protein hexamer. The six PII-like domains form two trimeric "lids" that cap the central cavity of the toroid on either side and provide only small openings to allow regulated entry of small molecules into the occluded chamber. The presence of the electron density of the bound ligand may provide important clues on the likely function of NIF3-like proteins.
Highlights
The protein encoded by the SA1388 gene from Staphylococcus aureus was chosen for structure determination to elucidate its domain organization and confirm our earlier remote homology based prediction that it housed a nitrogen regulatory PII protein-like domain
The PII domain is embedded within the central region of the polypeptide while the N- and C-terminal regions together belong to the NIF3 (NGG1p interacting factor 3)-like protein family
One subunit is traceable across the entire length of polypeptide from the N- to the C-terminus with the exception of a loop region containing 25 residues (168–193) in the central PII domain
Summary
The protein encoded by the SA1388 gene from Staphylococcus aureus was chosen for structure determination to elucidate its domain organization and confirm our earlier remote homology based prediction that it housed a nitrogen regulatory PII protein-like domain. Careful remote homology detection and manual analysis has in many occasions helped to glean useful structural and functional insights into these so-called "hypothetical proteins". Such studies involve a combination of profile based methods like transitive PSIBLAST [1], COMPASS [2] and HMMer [3] as well as structure prediction and fold recognition methods [4,5,6]. One such study [7] had analyzed sequences of the ubiquitously found protein modules homologous to the nitrogen regulatory PII proteins as defined in the COG [8] and Pfam databases [9] This comprehensive analysis expanded the PII protein superfamily to include five very divergent groups of proteins, with below random (~1%) pairwise sequence identities between some members of distant groups. The PII domain of these proteins would play some sort of ligand binding and signalling role analogous to that of classical PII proteins [10,11], while the function of the NIF3-like domains is not known
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