RESUMEN
Expression of the desC gene coding for acyl-lipid delta(9) desaturase of thermophilic cyanobacterium Synechocystis sp. PCC6803 was studied in Escherichia coli cells. In a hybrid gene constructed (desC-licBM3), a sequence of the native acyl-lipid delta(9) desaturase was fused in frame with the reporter gene coding for thermostable lichenase. Lichenase contained in the hybrid protein simplified selection and analysis of the expression of membrane desaturase in the heterologous host. Comparisons of the expression for the native and hybrid genes in bacterial cells showed that lichenase remained active and thermostable in the hybrid protein, while desaturase retains the capability of introducing a double bound in the corresponding position of fatty acids.
Asunto(s)
Proteínas Bacterianas/biosíntesis , Cianobacterias/enzimología , Escherichia coli/metabolismo , Ácido Graso Desaturasas/biosíntesis , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Bacterianas/genética , Clonación Molecular , Escherichia coli/genética , Ácido Graso Desaturasas/genética , Genes Reporteros , Glicósido Hidrolasas/genética , Proteínas Recombinantes de Fusión/genética , Estearoil-CoA DesaturasaRESUMEN
Coordination complexes of unsaturated rac-1,2-diacylglycerols (DAGs) with silver ions were separated by adsorption and reversed-phase TLC (Ag-TLC and Ag-rpTLC, respectively). During the Ag-TLC, the silver ion complexes were shown to be formed by the DAG coordination centers of various structures and only on the adsorbent surface. Separation of the complexes proceeds according to the adsorption mechanism, and there is an inverse exponential functional relationship between the DAG mobility and their double bond number. Meanwhile, during the Ag-rpTLC, the Ag(+)-complexes are formed only with double bonds, only in solution, and at a 1:1 ratio. The complexes are fractionated by partitioning between two liquid phases, and the relationship between the mobility and unsaturation of these complexes is directly proportional. Nevertheless, despite all the differences between the two TLC methods, the polarity of DAGs with a bent configuration of their acyl chains greatly exceeds that of DAGs with the same unsaturation but with the acyl-chain conformation close to extended: it is two to three times greater in Ag-TLC and 30-40% greater in Ag-rpTLC. In addition, the relationship between the mobility and unsaturation of DAG complexes exhibits quantitative rather than qualitative differences in both versions of argentation TLC. Thus, under all conditions of argentation liquid chromatography, the mobility of complexes and, therefore, their polarity are determined not only by their composition (unsaturation), but also by the spatial structure (conformation) of their molecules.