Role of the Periplasmic Chaperones Skp, SurA, and DegQ in Outer Membrane Protein Biogenesis in Neisseria meningitidis

Abstract

The periplasmic chaperones Skp, SurA, and DegP are implicated in the biogenesis of outer membrane proteins (OMPs) in Escherichia coli. Here, we investigated whether these chaperones exert similar functions in Neisseria meningitidis. Although N. meningitidis does not contain a homolog of the protease/chaperone DegP, it does possess a homolog of another E. coli protein, DegQ, which can functionally replace DegP when overproduced. Hence, we examined whether in N. meningitidis, DegQ acts as a functional homolog of DegP. Single skp, surA, and degQ mutants were easily obtained, showing that none of these chaperones is essential in N. meningitidis. Furthermore, all combinations of double mutants were generated and no synthetic lethality was observed. The absence of SurA or DegQ did not affect OMP biogenesis. In contrast, the absence of Skp resulted in severely lower levels of the porins PorA and PorB but not of other OMPs. These decreased levels were not due to proteolytic activity of DegQ, since porin levels remained low in a skp degQ double mutant, indicating that neisserial DegQ is not a functional homolog of E. coli DegP. The absence of Skp resulted in lower expression of the porB gene, as shown by using a P porB -lacZ fusion. We found no cross-species complementation when Skp of E. coli or N. meningitidis was heterologously expressed in skp mutants, indicating that Skp functions in a species-specific manner. Our results demonstrate an important role for Skp but not for SurA or DegQ in OMP biogenesis in N. meningitidis.

Distinct Roles of Secreted HtrA Proteases from Gram-negative Pathogens in Cleaving the Junctional Protein and Tumor Suppressor E-cadherin

“…S3A). Sequence alignment revealed a slightly higher identity of NgHtrA to the E. coli DegQ homolog (38%) than to DegP (37%) as recently suggested for Neisseria meningitidis (14). Hence, NgHtrA seems to be less related to other HtrAs, whereas HpHtrA and CjHtrA together with EpHtrA and SfHtrA exhibit high pairwise similarity (supplemental Fig.…”

“…Interestingly, NgHtrA did not cleave E-cadherin, which is explainable by the structural differences in the active pocket. NgHtrA was annotated as a serine protease, but obviously revealed a higher identity to the DegQ homolog consistent with HtrA from N. meningitidis (14), suggesting that DegQ and DegP functions are similar, but do not serve as general functional homologs (14). Among other differences, the exchanged Gln-263 crucially protrudes into the pocket of NgHtrA, which might interfere with E-cadherin binding and cleavage.…”

Involvement of Neisseria meningitidis Lipoprotein GNA2091 in the Assembly of a Subset of Outer Membrane Proteins

“…NspA is a small eight-stranded ␤-barrel OMP whose correctly folded form migrates more slowly in seminative SDS-PAGE than the denatured form (18). Using this assay, we found that all NspA was folded in the ⌬GNA2091 mutant (Fig.…”

“…The transformants were checked by PCR for the presence of the mutant allele using primer pair NMB2090-for and NMB2092-rev and for the absence of the wild-type allele using primer pair GNA2091-for1 and NMB2092-rev. Double skp::cat⌬GNA2091, ⌬surA⌬GNA2091 and ⌬bamC⌬GNA2091 mutants were obtained by introducing the ⌬GNA2091::kan allele into previously constructed ⌬surA::cat, ⌬bamC::cat, and insertional skp::cat mutants (11,18). For complementation purposes, GNA2091 was amplified by PCR using genomic DNA of HB-1 as template and primers GNA2091-for-NdeI and GNA2091-rev-AatII.…”

The LptD Chaperone LptE Is Not Directly Involved in Lipopolysaccharide Transport in Neisseria meningitidis

“…When neisserial cell envelopes are subjected to this procedure, LptD is detected as two high molecular weight (HMW) bands, indicating that the protein is present in complexes (32) (Fig. 5A).…”