Structural basis of pilus subunit recognition by the PapD chaperone.

The assembly of different types of virulence-associated surface fibers called pili in Gram-negative bacteria requires periplasmic chaperones. PapD is the prototype member of the periplasmic chaperone family, and the structural basis of its interactions with pilus subunits was investigated. Peptides corresponding to the carboxyl terminus of pilus subunits bound PapD and blocked the ability of PapD to bind to the pilus adhesin PapG in vitro. The crystal structure of PapD complexed to the PapG carboxyl-terminal peptide was determined to 3.0 A resolution. The peptide bound in an extended conformation with its carboxyl terminus anchored in the interdomain cleft of the chaperone via hydrogen bonds to invariant chaperone residues Arg8 and Lys112. Main chain hydrogen bonds and contacts between hydrophobic residues in the peptide and the chaperone stabilized the complex and may play a role in determining binding specificity. Site-directed mutations in Arg8 and Lys112 abolished the ability of PapD to bind pilus subunits and mediate pilus assembly in vivo, an indication that the PapD-peptide crystal structure is a reflection of at least part of the PapD-subunit interaction.

FimC is a periplasmic PapD-like chaperone that directs assembly of type 1 pili in bacteria.

Biogenesis of the type 1 pilus fiber in Escherichia coli requires the product of the fimC locus. We have demonstrated that FimC is a member of the periplasmic chaperone family. The deduced primary sequence of FimC shows a high degree of homology to PapD and fits well with the derived consensus sequence for periplasmic chaperones, predicting that it has an immunoglobulin-like topology. The chaperone activity of FimC was demonstrated by purifying a complex that FimC forms with the FimH adhesion. A fimC1 null allele could be complemented by the prototype member of the chaperone superfamily, PapD, resulting in the production of adhesive type 1 pili. The general mechanism of action of members of the chaperone superfamily was demonstrated by showing that the ability of PapD to assemble both P and type 1 pili was dependent on an invariant arginine residue (Arg-8), which forms part of a conserved subunit binding site in the cleft of PapD. We suggest that the conserved cleft is a subunit binding feature of all members of this protein family. These studies point out the general strategies used by Gram-negative bacteria to assemble adhesins into pilus fibers, allowing them to promote attachment to eukaryotic receptors.

Interactive surface in the PapD chaperone cleft is conserved in pilus chaperone superfamily and essential in subunit recognition and assembly.

The assembly of adhesive pili in Gram-negative bacteria is modulated by specialized periplasmic chaperone systems. PapD is the prototype member of the superfamily of periplasmic pilus chaperones. Previously, the alignment of chaperone sequences superimposed on the three dimensional structure of PapD revealed the presence of invariant, conserved and variable amino acids. Representative residues that protruded into the PapD cleft were targeted for site directed mutagenesis to investigate the pilus protein binding site of the chaperone. The ability of PapD to bind to fiber-forming pilus subunit proteins to prevent their participation in misassembly interactions depended on the invariant, solvent-exposed arginine-8 (R8) cleft residue. This residue was also essential for the interaction between PapD and a minor pilus adaptor protein. A mutation in the conserved methionine-172 (M172) cleft residue abolished PapD function when this mutant protein was expressed below a critical threshold level. In contrast, radical changes in the variable residue glutamic acid-167 (E167) had little or no effect on PapD function. These studies provide the first molecular details of how a periplasmic pilus chaperone binds to nascently translocated pilus subunits to guide their assembly into adhesive pili.

Respiratory health and lung function in 8-year-old children of very low birth weight: a cohort study.

In comparison with a cohort of normal birth weight children, those of very low birth weight (less than 1501 g birth weight) had more wheezing illnesses and hospital readmissions for respiratory problems in the first 2 years of life; from 2 years to 8 years of age respiratory health was unrelated to birth weight. Lung function measurements at 8 years of age in very low birth weight children were similar to expected values; few children had severely abnormal lung function. On univariate analyses, forced vital capacity (FVC) and forced expired volume in 1 second (FEV1), but not flow rates, were lower in children who had survived bronchopulmonary dysplasia. However, the univariate analyses were misleading, because bronchopulmonary dysplasia occurred more frequently with lower birth weight, and lower birth weight in turn was strongly related to reduced FVC and FEV1. After adjusting for birth weight and other potential confounding variables, FVC and FEV1 were unrelated to bronchopulmonary dysplasia, and to neonatal ventilation. Flow rates were largely uninfluenced by perinatal events, but were reduced in children with asthma or recurrent bronchitis at 8 years of age. Passive smoking was unrelated to lung function at 8 years of age. However, the effects of passive or active smoking, or perinatal events, on respiratory function or health beyond 8 years of age in very low birth weight survivors remain to be determined.

Free intra-peritoneal gas--an unusual aetiology.

The presence of free gas under one or both hemidiaphragms on an erect chest x-ray is most frequently the result of recent abdominal surgery or, in the absence of such surgery, is almost always indicative of a perforated abdominal viscus. A case is reported of a patient in whom pneumoperitoneum was discovered incidentally on a chest x-ray following extensive pelvic manipulation for acute inversion of the uterus.

CT appearance of a prosthetic methyl methacrylate mass mistaken for abscess.

A nonopaque methyl methacrylate mass that had the appearance of an abscess in a postoperative CT scan of the spine is described. Contained gas produced an attentuation range of +20 to -300 HU, with an average of -80 to -250 HU. This finding was confirmed by CT examination of a separate cement mass removed from a similar patient.

High-resolution CT scanning of the lumbar spine. Technique and anatomy.

Low back pain is a very common clinical problem, the cause of which is frequently poorly understood. Improvements in equipment design and scanning techniques have resulted in a better anatomical display of the lumbar spine by computed tomographic (CT) scanning. Advances in this area have provided a considerable insight into the causes of the lumbago-sciatica syndrome.