Coiled-coil structure of group A streptococcal M proteins. Different temperature stability of class A and C proteins by hydrophobic-nonhydrophobic amino acid substitutions at heptad positions a and d.
M proteins and M-like proteins, expressed on the surface of group A streptococci and binding to human plasma proteins, can be divided into two classes, A and C, depending on the structure of the central repeated regions. The class C proteins have been shown to be dimers with a coiled-coil structure. In this work, we have compared the structure and binding of a class A protein, Mrp4, and a class C protein, Arp4, expressed by the same bacterial strain. Circular dichroism spectra, gel filtration, and binding assays showed that both proteins had a coiled-coil dimer configuration and a high-affinity binding at 20 degrees C. However, striking differences were seen at 37 degrees C. The class A protein, Mrp4, was still a coiled-coil dimer with high affinity binding activity, whereas the class C protein, Arp4, had lost both the coiled-coil structure and binding activity. Raising the temperature even higher, Mrp4 retained the coiled-coil structure up to 70-90 degrees C. Furthermore, a recombinant protein, Mrp(C), in which the A-repeats of Mrp4 were replaced by the C-repeats of Arp4, lost its coiled-coil structure and fibrinogen-binding around 40-45 degrees C. These results suggest a high thermal stability of class A proteins and a low stability of class C proteins and that the structural basis for this can be found partly in the A- and C-repeats. Analysis of the amino acid sequences of the A- and C-repeats, revealed a large difference, 87% and 45%, respectively, in the content of hydrophobic amino acid residues in the positions regarded as important for the formation of the coiled-coil structure. In particular, several alanine residues in the A-repeats were replaced by serine residues in the C-repeats. Our results suggest that important structural and functional changes within the M protein family have evolved by specific hydrophobic-nonhydrophobic amino acid replacements.
Cedervall
T
T
Department of Cell and Molecular Biology, Lund University, Sweden.
Johansson
M U
MU
Akerström
B
B
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000942:Antigens, Bacterial / Q000737:chemistry*; D000954:Antigens, Surface / Q000737:chemistry; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D002352:Carrier Proteins / Q000737:chemistry*; D002850:Chromatography, Gel; D002942:Circular Dichroism; D005340:Fibrinogen / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000378:metabolism; D013297:Streptococcus pyogenes / Q000737:chemistry; D013329:Structure-Activity Relationship
D000596:Amino Acids; D000942:Antigens, Bacterial; D000954:Antigens, Surface; C062462:Arp4 protein, Streptococcus pyogenes; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D011994:Recombinant Proteins; C014689:streptococcal M protein; D005340:Fibrinogen