RESUMEN
Contrary to the accurate, hard-sphere depiction of monomeric hemoglobin in solution, sickle cell hemoglobin (HbS) polymerization/gelation requires attention to molecular interactions. From the temperature dependence of the osmotic compressibility of HbS gels, we were able to extract the entropy increase for concentrating HbS in this phase. Normalized per mole of water removed, the entropy increase from gel compression DeltaS(gel) is four times the previously measured DeltaS(trans), for the transition from monomeric HbS solution to HbS gel. The positive entropy change cannot emerge from the assembly of hard spheres but can indicate remodeling of HbS fibers driven by release of ordered water. The fourfold difference in DeltaS(gel) and DeltaS(trans) suggests that the act of initial fiber/gel formation from monomeric solution differs from the process of further polymerization due to tighter packing within the gel phase.
Asunto(s)
Hemoglobina Falciforme/química , Entropía , Geles , Humanos , Concentración Osmolar , Presión Osmótica , TermodinámicaRESUMEN
The structure of an "open state" of crystalline profilin:beta-actin has been solved to 2.65 A by X-ray crystallography. The open-state crystals, in 1.8 M potassium phosphate, have an expanded unit cell dimension in the c direction of 185.7 A compared with 171.9 A in the previously solved ammonium sulphate-stabilized "tight-state" structure. The unit cell change between the open and the tight states is accompanied by large subdomain movements in actin. Furthermore, the nucleotide in the open state is significantly more exposed to solvent, and local conformational changes in the hydrophobic pocket surrounding cysteine 374 occur during the transition to the tight state. Significant changes were observed at the N terminus and in the DNase-I binding loop. Neither the structure of profilin nor its contact with beta-actin are affected by the changes in the unit cell. Applying osmotic pressure to profilin:beta-actin crystals brings about a collapse of the unit cell comparable with that seen in the open to tight-state transition, enabling an estimate of the work required to cause this transformation of beta-actin in the crystals. The slight difference in energy between the open and collapsed states explains the extreme sensitivity of profilin:beta-actin crystals to changes in chemical and thermal environment.
Asunto(s)
Actinas/química , Actinas/metabolismo , Proteínas Contráctiles , Adenosina Trifosfato/metabolismo , Sitios de Unión , Cristalografía por Rayos X , Proteínas de Microfilamentos/química , Modelos Moleculares , Profilinas , Conformación Proteica , Sales (Química)/química , SolventesRESUMEN
Knowledge of the structure of actin in its various conformational states is important for understanding the diverse motile activities carried out by eukaryotic cells. Profilin:actin crystals provide a unique system for studying conformational states of actin, because they exhibit a high degree of polymorphism in response to environmental conditions while maintaining crystalline order. A preliminary comparison of two states of profilin:beta-actin crystals shows that crystal polymorphism involves movements of actin subdomains at hinge points homologous to those found in hexokinase, a protein whose polypeptide fold is related to actin. The homology of the hinge points in actin to those in hexokinase suggests that actin subdomain movements in profilin:beta-actin crystals have functional significance. We discuss how these movements could be related to structural transitions between states of filamentous actin in muscle contraction.
