Multiple folding states and disorder of ribosomal protein SA, a membrane receptor for laminin, anticarcinogens, and pathogens.

The human ribosomal protein SA (RPSA) is a multilocus protein, present in most cellular compartments. It is a multifunctional protein, which belongs to the ribosome but is also a membrane receptor for laminin, growth factors, prion, pathogenic microorganisms, toxins, and the anticarcinogen epigallocatechin gallate. It contributes to the crossing of the blood-brain barrier by neurotropic viruses and bacteria and is used as a biomarker of metastasis. RPSA includes an N-terminal domain, which is homologous to the prokaryotic ribosomal proteins S2, and a C-terminal extension, which is conserved in vertebrates. The structure of its N-domain has been determined from crystals grown at 17 °C. The structure of its C-domain remains unknown. We produced in Escherichia coli and purified the full-length RPSA and its N- and C-domains. We characterized the folding states of these recombinant proteins mainly by methods of fluorescence and circular dichroism spectrometry, in association with quantitative analyses of their unfolding equilibria, induced with heat or urea. The necessary equations were derived from first principles. The results showed that the N-domain unfolded according to a three-state equilibrium. The monomeric intermediate was predominant at the body temperature of 37 °C. It also existed in the full-length RPSA and bound ANS, a small fluorescent molecule. The C-domain was in an intrinsically disordered state. The recombinant N- and C-domains weakly interacted together. These results indicated a high plasticity of RPSA, which could be important for its multiple cellular localizations and functional interactions.

The folded and disordered domains of human ribosomal protein SA have both idiosyncratic and shared functions as membrane receptors.

The human RPSA [ribosomal protein SA; also known as LamR1(laminin receptor 1)] belongs to the ribosome but is also a membrane receptor for laminin, growth factors, prion, pathogens and the anticarcinogen EGCG (epigallocatechin-gallate). It contributes to the crossing of the blood-brain barrier by neurotropic viruses and bacteria, and is a biomarker of metastasis. RPSA includes an N-terminal domain, which is folded and homologous to the prokaryotic RPS2, and a C-terminal extension, which is intrinsically disordered and conserved in vertebrates. We used recombinant derivatives of RPSA and its N- and C-domains to quantify its interactions with ligands by in-vitro immunochemical and spectrofluorimetric methods. Both N- and C-domains bound laminin with K(D) (dissociation constants) of 300 nM. Heparin bound only to the N-domain and competed for binding to laminin with the negatively charged C-domain, which therefore mimicked heparin. EGCG bound only to the N-domain with a K(D) of 100 nM. Domain 3 of the envelope protein from yellow fever virus and serotypes-1 and -2 of dengue virus bound preferentially to the C-domain whereas that from West Nile virus bound only to the N-domain. Our quantitative in-vitro approach should help clarify the mechanisms of action of RPSA, and ultimately fight against cancer and infectious agents.