No abstract

Two overlapping promoters compete for RNA polymerase in the region that controls the expression of the galactose operon in Escherichia coli. Kinetics of open complex formation at P1 and P2 can be followed through the rate of formation of two specific abortive transcripts. The corresponding forward kinetic constants appear to be identical over a wide range of enzyme concentrations and temperatures, indicating that the two processes are strongly coupled. We propose a scheme accounting for our observations. In a first step, the competition between the two sites is a simple kinetic process, involving the "on" rate constants. In a second step, a slow reequilibration occurs, implicating the "off' rate constants and the conversion of one open complex to the other through a set of closed complexes. The first step is clearly affected when the complex between cyclic AMP and its receptor is bound at the activator site. An estimate of the various rate constants describing open complex formation at P1 and P2 is provided, as well as a qualitative description of the effect of the activator complex on these two pathways.

Polyacrylamide gel electrophoresis has been used to visualise and quantitate complexes between the Escherichia coli cyclic AMP receptor protein (CRP) and DNA fragments containing the promoter region of either the E. coli galactose or lactose operons. We show that, although CRP binding to the gal fragment is weaker than binding to the lac fragment, in each case, stable complexes are formed between one dimer of CRP and one molecule of DNA. We have examined the effects of a series of deletions and point mutations in the gal promoter region on CRP binding. From the position of deletions and mutations which prevent the formation of stable complexes, we deduce the location and extent of the sequence at the CRP binding site. We show that it covers approximately the same length of sequence as the binding site at the lac promoter. Unlike the lac site, the gal site contains no palindromic sequence. We discuss the importance of symmetry in the sequence at CRP binding sites and the validity of CRP binding consensus sequences which have been proposed.

We report the result of exposing silicone rubber to washed pig platelet suspensions that contained on average 0.018 mg of proteins/ml in solutions. This protein content is sufficiently low to reasonably neglect the protein adhesion to the material. On comparing the measured platelet adhesion from the platelet suspensions with that from blood, we find that when the gas nuclei normally present in the surface roughness of the material are removed the number of adhering platelets is the same. Thus, in the absence of the gas nuclei, the proteins in blood plasma play a negligible role in the platelet adhesion. In contrast, when both the gas nuclei and proteins are present, the maximum platelet adhesion was observed. From this and the above observation, it appears the gas nuclei affect one or more of the proteins, and this brings about an increased platelet adhesion. Finally, the platelet adhesion from the platelet suspensions was reduced after the removal of the gas nuclei. Thus the platelets themselves sense the change in the surface resulting from the removal of the gas nuclei.