Structure, Specificity, and Mode of Interaction for Bacterial Albumin-binding Modules

Abstract

We have determined the solution structure of an albumin binding domain of protein G, a surface protein of group C and G streptococci. We find that it folds into a left handed three-helix bundle similar to the albumin binding domain of protein PAB from Peptostreptococcus magnus. The two domains share 59% sequence identity, are thermally very stable, and bind to the same site on human serum albumin. The albumin binding site, the first determined for this structural motif known as the GA module, comprises residues spanning the first loop to the beginning of the third helix and includes the most conserved region of GA modules. The two GA modules have different affinities for albumin from different species, and their albumin binding patterns correspond directly to the host specificity of C/G streptococci and P. magnus, respectively. These studies of the evolution, structure, and binding properties of the GA module emphasize the power of bacterial adaptation and underline ecological and medical problems connected with the use of antibiotics.In the complex molecular interplay between a pathogen and its human host, protein-protein interactions play important roles. For instance, bacteria express surface proteins that interact with abundant human extracellular proteins with high affinity and specificity. In human plasma, albumin (HSA) 1 and immunoglobulins (Ig) are the quantitatively dominating proteins, and significant human pathogens have developed surface proteins that bind these and other plasma proteins (for references, see Ref 1). Two of the most well known such proteins are protein A of Staphylococcus aureus (2) and protein G of group C and G streptococci (3, 4), which both bind to the Fc region of IgG. In 1980, Myhre and Kronvall (5) reported that HSA could bind to the surface of various streptococcal species, including group C and G streptococci. It was later found that protein G was responsible also for the interaction with HSA (6). Protein G has separate binding domains for IgG and HSA (7,8), and as a result C and G streptococci are in vivo covered with an inner layer of IgG and an outer layer of HSA. Peptostreptococcus magnus are strictly anaerobic bacteria that are part of the indigenous human flora of the skin, oral cavity, and gastrointestinal and urogenital tracts. Some isolates of this species bind HSA (9), and notably these isolates are mostly from patients with deep wound infections (10), suggesting that HSA binding turns the commensal P. magnus into a potential pathogen. The surface protein of P. magnus binding HSA is called PAB, and it contains a domain of 45 residues showing a high degree of sequence homology with the HSA binding domains of protein G (11). Analysis of the gene encoding PAB suggested that this domain originates from protein G and has been transferred to and introduced into the pab gene through the action of a conjugational plasmid (related to pCF10 of Enterococcus faecalis) followed by a recombinational event (11). This interspecies exchange of a structurally well defined motif repre...

Biodistribution of a Bispecific Single-chain Diabody and Its Half-life Extended Derivatives

“…A prerequisite of the recycling of scDb-ABD via the FcRn is that the complex between scDb-ABD, albumin and FcRn remains stable in the acidic environment of the endosome. In QCM studies we found that the affinity of scDb-ABD is not decreased at pH 6.0 and that the measured affinities are similar to those determined by others for the binding of the single ABD domain to HSA at neutral pH (21)(22)(23).…”

Binding of Albumin Promotes Bacterial Survival at the Epithelial Surface

“…A3782). The GA module protein (residues 213-265 of protein PAB) was produced as described (31). Dermatan sulfate (DS) 36 and heparan sulfate (HS) 6 were provided by Lars-Åke Fransson.…”

Assessment of Genetic Shielding for Adenovirus Vectors

“…For sr39TK the NheI fragment was out of pShuttle.wt.E1-pIX-flag-TK [31]. The albumin binding domain 3 (ABD) from streptococcal Protein G (46 amino acid) [44,45] (Genbank X06173) was generated using two commercially synthesized single-stranded oligonucleotides (IDT), one starting at the 5' end, the other starting at the 3' end, with an overlapping central 15 nucleotide region. The oligonucleotides were annealed and extended, using Pfu Hifidelity TAQ (Stratagene), to form a double stranded (ds) cDNA of ABD with NheI restriction sites at both the 5' and 3' ends.…”

“…Pathogens use these proteins to avoid detection by the human immune system. The small ligand BAP has been successfully incorporated into pIX and been shown to conjugate to appropriately labeled ligands [30], validating our approach and therefore we decided to incorporate the albumin binding domain 3 from Streptococcal Protein G (ABD) [44,45] into pIX to be utilized as a docking site for albumin.…”

“…1). Bacteria express surface proteins that interact with human extracellular proteins including several natural albumin binding proteins that are well characterized, such as motifs in Protein G and Protein PAB [44]. Pathogens use these proteins to avoid detection by the human immune system.…”