Crystallization and preliminary X-ray diffraction studies of two domains of a bilobed extra-cytoplasmic function sigma factor SigC from Mycobacterium tuberculosis

Abstract

Sigma factors are transcription-regulatory proteins that bind to RNA polymerase and facilitate promoter recognition. The so-called extracytoplasmic function sigma factors help a bacterium to respond to environmental conditions. Mycobacterium tuberculosis SigC (σ^C) is an extracytoplasmic sigma factor that is essential for lethality in a mouse model of infection and is conserved in all pathogenic mycobacterial species. This protein consists of two domains that are connected by an ~25-amino-acid linker. The N-terminal domain contains the σ₂ DNA-binding motif, whereas the σ₄ motif is located in the C-terminal domain. Native σ^C did not yield diffraction-quality crystals. However, two of its domains have been cloned, expressed and crystallized: σ^C₂ (12.3 kDa) and σ^C₄ (7.5 kDa). The σ^C₂ crystals belong to the hexagonal space group P6₁, with unit-cell parameters a = b = 85.28, c = 79.63 Å, and native X-ray diffraction data were collected from this domain to 2.7 Å on an in-house X-ray home source. The σ^C₄ crystals belong to the cubic space group F23, with unit-cell parameters a = b = c = 161.21 Å. X-ray diffraction data were collected from this domain to 3.1 Å, also on an in-house X-ray source.