Crystallization and preliminary X-ray diffraction study of an idiotope-anti-idiotope Fv-Fv complex.
A complex between the Fv fragment of an anti-hen eggwhite lysozyme antibody (D1.3) and the Fv fragment of an antibody specific for an idiotypic determinant of D1.3 has been crystallized in a form suitable for X-ray diffraction analysis. Both Fv fragments were expressed in soluble form in Escherichia coli and purified by affinity chromatography; diffraction-quality crystals were only obtained following separation of each Fv into distinct isoelectric forms. The crystals belong to space group C2, have unit cell dimensions a = 152.8 A, b = 79.4 A, c = 51.5 A, beta = 100.2 degrees, and diffract to better than 2.2 A resolution. The solvent content of the crystals is approximately 60% (v/v) with one Fv-Fv complex in the asymmetric unit. The ability to readily express both components of an antigen-antibody system in bacteria will allow us to rigorously assess the energetic contribution of individual amino acids to complex formation through pairwise mutagenesis of interacting residues.
Goldbaum
F A
FA
Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute, Rockville 20850.
Fields
B A
BA
Cauerhff
A
A
Ysern
X
X
Houdusse
A
A
Eisele
J L
JL
Poljak
R J
RJ
Mariuzza
R A
RA
D000595:Amino Acid Sequence; D000888:Antibodies, Anti-Idiotypic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000936:Antigen-Antibody Complex / Q000737:chemistry*; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003460:Crystallization; D018360:Crystallography, X-Ray; D005803:Genes, Immunoglobulin / Q000235:genetics; D007128:Immunoglobulin Fragments / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D009113:Muramidase
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; C055966:immunoglobulin Fv; D009113:Muramidase