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

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T1 - 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

AU - Cedervall, T

AU - Johansson, M U

AU - Akerström, B

PY - 1997/4/22

Y1 - 1997/4/22

N2 - 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.

AB - 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.

KW - Amino Acid Sequence

KW - Amino Acids/analysis

KW - Antigens, Bacterial/chemistry

KW - Antigens, Surface/chemistry

KW - Bacterial Outer Membrane Proteins

KW - Bacterial Proteins/chemistry

KW - Carrier Proteins/chemistry

KW - Chromatography, Gel

KW - Circular Dichroism

KW - Fibrinogen/metabolism

KW - Hot Temperature

KW - Humans

KW - Molecular Sequence Data

KW - Mutagenesis, Site-Directed

KW - Protein Conformation

KW - Protein Structure, Secondary

KW - Recombinant Proteins/metabolism

KW - Streptococcus pyogenes/chemistry

KW - Structure-Activity Relationship

U2 - 10.1021/bi962971q

DO - 10.1021/bi962971q

M3 - Article

VL - 36

SP - 4987

EP - 4994

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 16

ER -