title: On the interaction of alpha-crystallin with unfolded proteins.
abstract: alpha-Crystallin, a major protein component of the lens, has chaperone-like properties whereby it prevents destabilised proteins from precipitating out of solution. It does so by forming a soluble high-molecular-weight (HMW) complex. A spectroscopic investigation of the HMW complex formed between a variety of unfolded proteins and bovine alpha-crystallin is presented in this paper. As monitored by fluorescence spectroscopy, a large amount of the hydrophobic probe, 8-anilino-1-naphthalene sulfonate (ANS) binds to the HMW complex implying that the complexed proteins (alcohol dehydrogenase (ADH), gamma-crystallin and rhodanese) are bound in an unfolded, possibly molten-globule state. The interaction between the anionic surfactant, sodium dodecyl sulfate (SDS) and ADH at high temperatures gives rise to a similar large increase in ANS fluorescence to that for the complex between alpha-crystallin and ADH. SDS, like alpha-crystallin, therefore complexes to proteins in their unfolded state leaving a large hydrophobic surface exposed to solvent. Unlike other chaperones (e.g., GroEL, DnaK and SecB), alpha-crystallin does not interact with unfolded, hydrophobic but stable proteins (e.g., reduced and carboxymethylated alpha-lactalbumin and alpha-casein). It is concluded that alpha-crystallin will only complex with proteins that are about to precipitate out of solution, i.e., ones that are severely compromised. 1H-NMR spectroscopy of the HMW complex formed between alpha-crystallin and gamma-crystallin indicates that the short C-terminal extension of alpha B-crystallin, but not that of alpha A-crystallin, has lost its flexibility in the complex implying that the former is involved in interactions with the unfolded gamma-crystallin molecule, possibly electrostatically via its two C-terminal lysine residues.
authors.affiliation: Australian Cataract Research Foundation, Department of Chemistry, University of Wollongong, NSW, Australia.
authors.forename: J A
authors.initials: JA
authors.lastname: Carver
authors.forename: N
authors.initials: N
authors.lastname: Guerreiro
authors.forename: K A
authors.initials: KA
authors.lastname: Nicholls
authors.forename: R J
authors.initials: RJ
authors.lastname: Truscott
mesh_terms: D000426:Alcohol Dehydrogenase / Q000737:chemistry; D000817:Anilino Naphthalenesulfonates; D000818:Animals; D002417:Cattle; D003459:Crystallins / Q000737:chemistry* / Q000302:isolation & purification; D006358:Hot Temperature; D007908:Lens, Crystalline / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D018832:Molecular Chaperones / Q000737:chemistry; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry*; D012967:Sodium Dodecyl Sulfate; D012996:Solutions; D013050:Spectrometry, Fluorescence
chemical_list: D000817:Anilino Naphthalenesulfonates; D003459:Crystallins; D018832:Molecular Chaperones; D011506:Proteins; D012996:Solutions; D012967:Sodium Dodecyl Sulfate; C035763:5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate); D000426:Alcohol Dehydrogenase