KapB is a lipoprotein required for KinB signal transduction and activation of the phosphorelay to sporulation in Bacillus subtilis

Abstract

SummaryKinB is one of the two major histidine kinases that provide phosphate input in the phosphorelay to produce Spo0AϳP, the key transcription factor controlling the initiation of sporulation. A search for insertion mutants affected in activation of KinB-dependent sporulation led to the identification of the lgt locus encoding the lipoprotein glyceryltransferase required for the lipid modification of prolipoproteins before their cleavage and translocation across the cytoplasmic membrane. In parallel, a putative lipoprotein signal peptide cleavage site was detected in KapB, known to be strictly required for KinB-mediated sporulation and located downstream of KinB in a single transcription unit. Using PhoA peptide fusions, we have shown that KapB signal-peptide can direct active alkaline phosphatase to the outer surface of the cytoplasmic membrane in an LGT-dependent manner, strongly suggesting that KapB is a lipoprotein tethered to the outer face of the cytoplasmic membrane via a lipid anchor. As KapB proved to be dispensable for expression of the kinBkapB operon, a chimeric kinase was built consisting of KinA sensor domain fused to KinB kinase domain (KinAЈ-ЈB) to assess (i) the involvement of KapB in catalysis of the kinase reaction, and (ii) the ability of KinB to phosphorylate Spo0F in vitro. It was shown that KapB is dispensable for both in vivo and in vitro activation of the phosphorelay by the KinAЈ-ЈB chimera and that KinAЈ-ЈB phosphor ylates Spo0F directly in vitro. Models for the role of KapB in regulating KinB activity are discussed.

Role of anionic phospholipids in the adaptation of Bacillus subtilis to high salinity

“…One possible explanation for the major variations in lipids occurring during stress or growth phase is that they would influence the membrane structure, thus affecting the activity of enzymes involved in different processes such as sporulation (Dartois et al, 1997), osmosensing (Poolman et al, 2004) and modifications of protein function (Jensen & Mouritsen, 2004). van der Heide et al (2001), while studying the ionic regulation of the OpuA transporter of Lactococcus lactis in proteoliposomes containing different phospholipid composition, showed that only the charge present on the lipid headgroups influences the osmotic activation profile of this transporter.…”

Lipoprotein biogenesis in Gram-positive bacteria: knowing when to hold ‘em, knowing when to fold ‘em

“…In addition to the sensing of peptides described earlier, lipoproteins have roles in cell envelope sensing processes, including the modulation of two-component signal transduction systems [44,69,70] and the Bacillus germinant receptors GerAC, GerBC, GerD and GerKC [46,71]. Moreover, genome analyses have consistently identified putative lipoproteins predicted to have important roles in cell envelope stability and cell wall cross-linking or remodelling such as penicillin binding proteins and peptidoglycan hydrolases [18,54].…”

“…Unsurprisingly, given the variety of functions attributable to lipoproteins, lgt or lsp mutations have pleiotropic effects. For example, lgt mutants of B. subtilis exhibit defects in cytochrome caa 3 activity [43], protein secretion [33], germination and sporulation [44][45][46] that can be correlated to impaired functions of specific lipoproteins.…”

Phosphorylation of Spo0A by the Histidine Kinase KinD Requires the Lipoprotein Med in Bacillus subtilis

“…Although there is no evidence of a direct interaction between KinB and KapB, the fact that their genes are in the same operon has led to the idea that the two proteins physically interact (10). Med acts by stimulating phosphorylation (and under some conditions, dephosphorylation) of Spo0A indirectly through KinD.…”

“…To be functional, KinB also requires the presence of a lipoprotein, KapB. The kapB gene is in an operon with the kinB gene, and the two proteins are thought to form a complex in the cytoplasmic membrane (10).…”