Monoclonal antibodies for the structural analysis of the Na+/H+ antiporter NhaA from Escherichia coli.
Since their advent some 25 years ago, monoclonal antibodies have developed into powerful tools for structural and functional analysis of their cognate antigens. Together with the respective antigen binding fragments, antibodies offer exclusive capacities in detection, characterization, purification and functional assays for every given ligand. Antibody-fragment mediated crystallization represents a major advance in determining the three-dimensional structure of membrane-bound protein complexes. In this review, we focus on the methods used to generate monoclonal antibodies against the NhaA antiporter from Escherichia coli as a paradigm of secondary transporters. We describe examples on how antibodies are helpful in understanding structure and function relationships for this important class of integral membrane proteins. The generated conformation-specific antibody fragments are highly valuable reagents for co-crystallization attempts and structure determination of the antiporter.
Venturi
Miro
M
Structural Biology Section, Vaccine Research Center/National Institutes of Health, Bethesda, MD 20892, USA.
Hunte
Carola
C
D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D000915:Antibody Affinity; D003001:Cloning, Molecular; D003460:Crystallization; D000939:Epitopes / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry; D007128:Immunoglobulin Fragments / Q000737:chemistry; D007700:Kinetics; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D011487:Protein Conformation; D017923:Sodium-Hydrogen Exchangers / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D000939:Epitopes; D029968:Escherichia coli Proteins; D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D008565:Membrane Proteins; C102839:NhaA protein, E coli; D017923:Sodium-Hydrogen Exchangers