Structural stabilization displayed in the soluble phase of cocrystallization of flavianic acid with trypsin in acid media.
Coprecipitation and cocrystallization of proteins with synthetic dyes are known to involve reversible denaturation processes which can offer specificity towards a target protein. Although the knowledge of conformational equilibrium and how to control it are central into the basic molecular dynamics of protein precipitation, the exact molecular mechanism of the precipitation remains unknown. Aiming at understanding the events that take place before the coprecipitation step of generic dye-protein systems, we investigated the binding of flavianic acid to bovine trypsin, using approaches of visible and second-derivative ultraviolet spectroscopy, viscosimetry, densimetry and circular dichroism. The results suggest a restricted transconformation of the macromolecule linked to dye binding at a stoichiometry of 1:1. An increase on the protein secondary structures occurred together with an electro-constriction effect on trypsin, a borderline event to the coprecipitation process, suggesting a stabilized structure for trypsin as a ligand-induced molten globule-like state.
Schneedorf
J M
JM
Lab. Fitofármacos, Faculdade de Medicina, Universidade de Alfenas, Rod. MG 179, Km 0, Alfenas, Minas Gerais, CEP 37130-000, Brazil.
Santoro
M M
MM
Mares-Guia
M
M
D000143:Acids / Q000378:metabolism; D002942:Circular Dichroism; D004396:Coloring Agents / Q000737:chemistry / Q000378:metabolism*; D003460:Crystallization; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D008024:Ligands; D008958:Models, Molecular; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012995:Solubility; D013056:Spectrophotometry, Ultraviolet; D013451:Sulfonic Acids / Q000737:chemistry / Q000378:metabolism*; D017186:Titrimetry; D014357:Trypsin / Q000737:chemistry* / Q000378:metabolism*; D014783:Viscosity
D000143:Acids; D004396:Coloring Agents; D008024:Ligands; D013451:Sulfonic Acids; C438653:flavianic acid; D014357:Trypsin