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Molecular structure of the lipoamide dehydrogenase domain of a surface antigen from Neisseria meningitidis
I Li de la Sierra1, L Pernot, T Prangé
1CIGB, Cubanacan, La Habana, Cuba.
Abstract:
The protein p64k from the surface of the Neisseria meningitidis bacteria has been characterized as a two-domain protein. It contains a dihydrolipoamide dehydrogenase domain of 482 residues, involving a FAD prosthetic group as a cofactor, and a smaller lipoic acid binding domain of 86 residues. The two domains are joined by a flexible segment rich in alanine and proline residues. The structure of the dihydrolipoamide dehydrogenase domain was determined by X-ray diffraction. It was solved by a combination of molecular replacement and multiple isomorphous replacement techniques and refined to 2.7 A resolution. In the crystal, the recombinant p64k mimics the functional homo-dimer by using one of the crystallographic 2-fold axes. The reactive disulphide bridge Cys161-Cys166 is in the oxidised state and the FAD is bound in an extended conformation. This main domain contains the major antigenic determinant of the protein, an extended loop of 32 residues at the surface of the protein. A mis-attribution at residue 553 in the sequence has been detected by inspection of electron density maps and the geometry. However, when compared to the other dihydrolipoamide dehydrogenases, there are some significant differences: (1) an unusual number of cis-proline residues and (2) a new motif built around a 2-fold axis by the sulphur atoms of residues Met558, Cys560 and their symmetry related equivalents.
Insights
The Neisseria meningitidis p64k protein has two domains: a dihydrolipoamide dehydrogenase domain and a lipoic acid binding domain. Its crystal structure reveals key features, including the antigenic determinant loop and unique structural motifs.
Area of Science:
- Bacteriology
- Structural Biology
- Biochemistry
Background:
- Neisseria meningitidis is a significant human pathogen.
- The p64k protein is a surface protein of N. meningitidis.
- p64k is a two-domain protein with potential functional significance.
Purpose of the Study:
- To characterize the structure of the p64k protein from Neisseria meningitidis.
- To elucidate the structural basis of p64k's function and antigenicity.
- To identify unique structural features compared to other dihydrolipoamide dehydrogenases.
Main Methods:
- X-ray diffraction was used to determine the structure of the dihydrolipoamide dehydrogenase domain.
- Molecular replacement and multiple isomorphous replacement techniques were employed for structure solution.
- Structure refinement was performed to 2.7 A resolution.
Main Results:
- The crystal structure of the p64k dihydrolipoamide dehydrogenase domain was solved.
- The recombinant protein formed a functional homo-dimer in the crystal.
- The major antigenic determinant was identified as a surface-exposed loop.
- Unique structural features, including cis-proline residues and a novel sulfur-based motif, were observed.
Conclusions:
- The determined structure provides insights into the molecular organization of p64k.
- The findings highlight unique structural adaptations in this bacterial protein.
- The structure aids in understanding p64k's role in N. meningitidis pathogenesis and potential as a vaccine target.