Environmental properties of long-chain alcohols. Part 2: Structure-activity relationship for chronic aquatic toxicity of long-chain alcohols.

Daphnia magna reproduction tests were performed with C(10), C(12), C(14) and C(15) alcohols to establish a structure-activity relationship of chronic effects of long-chain alcohols. The data generation involved substantial methodological efforts due to the exceptionally rapid biodegradability of the test substances and the need to test as close as possible to their water solubility limits. Test concentrations were determined by GC-MS before and after test solution renewal. Whereas apparent toxicity based on survival and reproduction increased with increasing C-chain lengths up to C(14), observations of toxicity to C(15) alcohol were not in line with lower chain lengths due to the lack of toxicity below the level of water solubility. When omitting C(15), the slope of most (Q)SARs approach -1, being consistent with the expectation of a non-polar narcotic mode of action. Further testing at higher chain lengths is not sensible due to progressively lower solubility, at remaining biodegradability. Effects on mortality and reproduction are not expected below the level of water solubility.

An outer membrane protein (OmpA) of Escherichia coli K-12 undergoes a conformational change during export.

Pulse-chase experiments were performed to follow the export of the Escherichia coli outer membrane protein OmpA. Besides the pro-OmpA protein, which carries a 21-residue signal sequence, three species of ompA gene products were distinguishable. One probably represented an incomplete nascent chain, another the mature protein in the outer membrane, and the third, designated imp-OmpA (immature processed), a protein which was already processed but apparently was still associated with the plasma membrane. The pro- and imp-OmpA proteins could be characterized more fully by using a strain overproducing the ompA gene products; pro- and imp-OmpA accumulated in large amounts. It could be shown that the imp- and pro-OmpA proteins differ markedly in conformation from the OmpA protein. The imp-OmpA, but not the pro-OmpA, underwent a conformational change and gained phage receptor activity upon addition of lipopolysaccharide. Utilizing a difference in detergent solubility between the two polypeptides and employing immunoelectron microscopy, it could be demonstrated that the pro-OmpA protein accumulated in the cytoplasm while the imp-OmpA was present in the periplasmic space. The results suggest that the pro-OmpA protein, bound to the plasma membrane, is processed, and the resulting imp-OmpA, still associated with the plasma membrane, recognizes the lipid A moiety of the lipopolysaccharide. The resulting conformational change may then force the protein into the outer membrane.

Rate of translation and kinetics of processing of newly synthesized molecules of two major outer-membrane proteins, the OmpA and OmpF proteins, of Escherichia coli K12.

The rate of synthesis of the OmpA and OmpF proteins, two of the major outer membrane proteins of Escherichia coli K12, was determined. At 25 degrees C both proteins were translated at 6.5 amino acids/s, and the OmpF protein was translated at 15 amino acids/s at 37 degrees C. The former rate corresponded to a synthesis time of just over 50 s for both proteins, which is significantly faster than their reported rates of assembly into the outer membrane at 25 degrees C. The kinetics of processing of the pro-OmpF protein were also investigated in detail, and the pro-OmpF half-life estimated to be 3-5 s at 25 degrees C. However a fraction of the precursor was processed more slowly, which may explain the discrepancy between these data and our earlier published estimate of 30 s. Pro-OmpA protein was processed with similar kinetics. These results demonstrate that the rate-limiting step in the assembly of both proteins into the outer membrane is post-translational and follows the processing step.

Precursor proteins are intermediates in vivo in the synthesis of two major outer membrane proteins, the OmpA and OmpF proteins, of Escherichia coli K12.

The OmpA and OmpF proteins are major outer membrane proteins of Escherichia coli K12. Their precursors, the pro-OmpA and pro-OmpF proteins, have been detected in vivo in pulse-labelling experiments carried out with [35S]methionine at 25 degrees C. Wehn the pulse was at 37 degrees C, however, no precursors were detected. The pulse-labelled precursors were processed rapidly and quantitatively into mature protein at 25 degrees C. The apparent half-life of the pro-OmpF protein was estimated to be 30 s, and the pro-OmpA protein may be processed even faster. In short pulses (10 s) the precursors of both proteins were the predominant labelled species, indicating that at 25 degrees C processing does not start until chain elongation of the precursor is almost, if not entirely, complete. When French press lysates of cells pulse-labelled for 10 s were subjected to sucrose gradient centrifugation to separate the inner and outer membranes, both precursors comigrated with the inner membrane.

Major proteins of the outer cell envelope membrane of Escherichia coli K12: multiple species of protein I differ in primary structure.

Protein I, one of the major outer membrane proteins of E. coli in most K12 strains is represented by two very similar polypeptides Ia and Ib. Sequential mutations (involving selections for phage resistance) can lead to loss of proteins Ia and Ib. Among "revertants" of such Ia-Ib- mutants clones exist that instead of Ia or Ib produce a third species of protein I, polypeptide Ic. Ichihara and Mizushima [J. Biochem. 83, 1095--1100 (1978)] have shown that proteins Ia and Ib exhibit differences in primary structure. Here evidence is presented indicating that protein Ic also is not identical in primary structure with Ia or Ib. Thus, 3 very similar structural genes appear to exist for the protein I species known to date, and that for Ic normally is silent. Introduction of a functional Ic locus into a Ia+ Ib+ strain caused expression of all three proteins with a reduced rate of synthesis of protein Ia.