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Crystal structure of Klebsiella pneumoniae sigma4 of sigmaS fusing with the RNA polymerase beta-flap-tip-helix in complex with -35 element DNA
Structural highlights
FunctionRPOB_KLEP7 DNA-dependent RNA polymerase catalyzes the transcription of DNA into RNA using the four ribonucleoside triphosphates as substrates.[HAMAP-Rule:MF_01321]RPOS_ECOLI Sigma factors are initiation factors that promote the attachment of RNA polymerase to specific initiation sites and are then released. This sigma factor is the master transcriptional regulator of the stationary phase and the general stress response. Controls, positively or negatively, the expression of several hundred genes, which are mainly involved in metabolism, transport, regulation and stress management.[HAMAP-Rule:MF_00959][1] [2] [3] [4] [5] Protects stationary phase cells from killing induced by endoribonuclease MazF.[6] Publication Abstract from PubMedIn class II transcription activation, the transcription factor normally binds to the promoter near the -35 position and contacts the domain 4 of sigma factors (sigma4 ) to activate transcription. However, sigma4 of sigma(70) appears to be poorly folded on its own. Here, by fusing sigma4 with the RNA polymerase beta-flap-tip-helix, we constructed two sigma4 chimera proteins, one from sigma(70) sigma 4 70 c and another from sigma(S) sigma 4 S c of Klebsiella pneumoniae. The two chimera proteins well folded into a monomeric form with strong binding affinities for -35 element DNA. Determining the crystal structure of sigma 4 S c in complex with -35 element DNA revealed that sigma 4 S c adopts a similar structure as sigma4 in the Escherichia coli RNA polymerase sigma(S) holoenzyme and recognizes -35 element DNA specifically by several conserved residues from the helix-turn-helix motif. By using nuclear magnetic resonance (NMR), sigma 4 70 c was demonstrated to recognize -35 element DNA similar to sigma 4 S c . Carr-Purcell-Meiboom-Gill relaxation dispersion analyses showed that the N-terminal helix and the beta-flap-tip-helix of sigma 4 70 c have a concurrent transient alpha-helical structure and DNA binding reduced the slow dynamics on sigma 4 70 c . Finally, only sigma 4 70 c was shown to interact with the response regulator PmrA and its promoter DNA. The chimera proteins are capable of -35 element DNA recognition and can be used for study with transcription factors or other factors that interact with domain 4 of sigma factors. Structural basis for -35 element recognition by sigma4 chimera proteins and their interactions with PmrA response regulator.,Lou YC, Chou CC, Yeh HH, Chien CY, Sadotra S, Hsu CH, Chen C Proteins. 2020 Jan;88(1):69-81. doi: 10.1002/prot.25768. Epub 2019 Jul 22. PMID:31293000[7] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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