Shot-gun phage display mapping of two streptococcal cell-surface proteins.
We have used a phage display shot-gun cloning technique to map the binding domains in two cell surface proteins from animal group C streptococci. The proteins, MAG and ZAG, have affinity for alpha (2)-macroglobulin (alpha (2)M), serum albumin and IgG. In this work, parts of cloned i mag and zag genes were randomly cloned into a phagemid vector, and recombinant phages expressing alpha (2)-M- or albumin-binding activity were isolated through panning against immobilized alpha (2)M or albumin. Analysis of the clones revealed two distinct alpha (2)M-binding sites in protein MAG and two slightly overlapping binding sites in protein ZAG. The minimal albumin-binding domain in protein ZAG, as deduced from the affinity selected clones, consisted of 42 amino acids. These results show that the phage display shot-gun cloning is a rapid and convenient way to characterize the binding site(s) in receptor proteins without any prior knowledge of their number, size, and localization.
Jacobsson
K
K
Department of Microbiology, Swedish University of Agricultural Sciences, Uppsala, Sweden.
Jonsson
H
H
Lindmark
H
H
Guss
B
B
Lindberg
M
M
Frykberg
L
L
D000418:Albumins / Q000378:metabolism*; D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003001:Cloning, Molecular / Q000379:methods; D008969:Molecular Sequence Data; D019151:Peptide Library; D011485:Protein Binding; D013291:Streptococcus / Q000235:genetics*; D000511:alpha-Macroglobulins / Q000378:metabolism*
D000418:Albumins; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; C087500:MAG protein, Streptococcal; D019151:Peptide Library; C108290:ZAG protein, Streptococcal; D000511:alpha-Macroglobulins