title: C repeats of the streptococcal M1 protein achieve the human serum albumin binding ability by flanking regions which stabilize the coiled-coil conformation.
abstract: The M and M-like proteins of Streptococcus pyogenes are fibrous cell surface proteins. They have multiple binding sites for several human proteins and are composed of the C-terminal anchor domain, the alpha-helical coiled-coil domain, and the N-terminal non-coiled-coil domain. The coiled-coil domain of the M1 protein consists of repeat units called B, C, and D and a spacer unit S between B and C. Recombinant fragments A-B-S-C-D, A-B-S, B-S-C, S-C, S-C-D, C-D, and C of the coiled-coil domain were studied by analyzing their secondary structures and binding affinities to human serum albumin (HSA). As shown by circular dichroism, all fragments are in an alpha-helical conformation. C-D and S-C-D form coiled coils at room temperature and bind below 37 degrees C with high affinity to HSA. C-D and S-C-D unfold in two steps with Tm values of approximately 31 and approximately 65 degrees C; complex formation with HSA increases the unfolding temperatures. B-S-C has a lower alpha-helical content, a less pronounced coiled-coil conformation, and a reduced thermal stability, binds HSA weaker, and is only slightly stabilized by HSA binding in comparison to C-D and S-C-D. C and S-C are less stable than the other fragments and are not organized as coiled coils showing some features of alpha-helical single strands only below 20 degrees C, and binding of HSA was not observed. The results indicate that the formation of coiled-coil structures, supported by flanking D regions and, to a lesser extent also B regions, is essential for the binding of C repeat units to HSA.
authors.affiliation: Institute of Experimental Microbiology, University of Jena, Winzerlaer Strasse 10, D-07745, Jena, Germany.
authors.forename: K
authors.initials: K
authors.lastname: Gubbe
authors.forename: R
authors.initials: R
authors.lastname: Misselwitz
authors.forename: K
authors.initials: K
authors.lastname: Welfle
authors.forename: W
authors.initials: W
authors.lastname: Reichardt
authors.forename: K H
authors.initials: KH
authors.lastname: Schmidt
authors.forename: H
authors.initials: H
authors.lastname: Welfle
mesh_terms: D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D002850:Chromatography, Gel; D002942:Circular Dichroism; D006801:Humans; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012091:Repetitive Sequences, Nucleic Acid; D012709:Serum Albumin / Q000378:metabolism* / Q000494:pharmacology; D013297:Streptococcus pyogenes / Q000378:metabolism; D013696:Temperature; D013816:Thermodynamics
chemical_list: D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D010446:Peptide Fragments; D012709:Serum Albumin; C014689:streptococcal M protein