RESUMEN
We show that the cAMP receptor protein (Crp) binds to DNA as several different conformers. This situation has precluded discovering a high correlation between any sequence property and binding affinity for proteins that bend DNA. Experimentally quantified affinities of Synechocystis sp. PCC 6803 cAMP receptor protein (SyCrp1), the Escherichia coli Crp (EcCrp, also CAP) and DNA were analyzed to mathematically describe, and make human-readable, the relationship of DNA sequence and binding affinity in a given system. Here, sequence logos and weight matrices were built to model SyCrp1 binding sequences. Comparing the weight matrix model to binding affinity revealed several distinct binding conformations. These Crp/DNA conformations were asymmetrical (non-palindromic).
Asunto(s)
Proteínas Bacterianas/química , Proteína Receptora de AMP Cíclico/química , ADN/química , Algoritmos , Proteínas Bacterianas/metabolismo , Sitios de Unión , Proteína Receptora de AMP Cíclico/metabolismo , Proteína Receptora de AMP Cíclico/normas , ADN/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Modelos Biológicos , Conformación de Ácido Nucleico , Regiones Promotoras Genéticas , Unión Proteica , Conformación Proteica , Estándares de Referencia , Análisis de Secuencia de ADN , SynechocystisRESUMEN
The cAMP receptor protein (Crp) is a global transcriptional regulator that binds sequence-specific promoter elements when associated with cAMP. In the motile cyanobacterium Synechocystis sp. strain PCC 6803, intracellular cAMP increases when dark-adapted cells are illuminated. Previous work has established that Crp binds proposed Crp target sites upstream of slr1351 (murF), sll1874 (chlA(II)), sll1708 (narL), slr0442 and sll1268 in vitro, and that slr0442 is downregulated in a crp mutant during photoautotrophic growth. To identify additional Crp target genes in Synechocystis, 11 different Crp binding sites proposed during a previous computational survey were tested for in vitro sequence-specific binding and crp-dependent transcription. The results indicate that murF, chlA(II) and slr0442 can be added as 'target genes of Sycrp1' in Synechocystis. Promoter mapping of the targets revealed the same close association of RNA polymerase and Crp as that found in Escherichia coli class I and class II Crp-regulated promoters, thereby strongly suggesting similar mechanisms of transcriptional activation.
