[Substitution treatment of drug-dependent patients]. / Substitutionsbehandlung Drogenabhängiger.

The author discusses the network of acts of parliament, subordinate legislation and decisions of the courts. Even if a substitution is legal, this does not necessarily mean that the expenses are covered by the statutory medical insurances. While in certain cases the insurances bear the costs of a treatment with methadone, the expenses for codeine and dihydrocodeine can be covered in no case. But it is possible to prescribe both at the expense of the patient. According to the law and the opinions of leading experts it is obligatory that the distribution of substitutes be combined with further medical, psychological and social care. For the latter the insurances cannot be charged. The essay concludes with presumptions about the future development of the regulations in force.

A stochastic description of evolutionary processes in underoccupied systems.

A stochastic description for underoccupied systems (large networks with small overall particle number) is given and represented by graphs. Evolutionary processes on this network are considered, especially the mutation process between components (hopping from one component to another) and the selection process.

Degrees of relatedness of T-even type E. coli phages using different or the same receptors and topology of serologically cross-reacting sites.

The relatedness of a series of T-even like phages which use the Escherichia coli outer membrane protein OmpA as a receptor, and the classical phages T2, T4 and T6 has been investigated. Immunoelectron microscopy and the pattern of phage resistance in bacterial mutants revealed that: (i) phages of this morphology do not necessarily cross-react serologically; (ii) phages using different receptors may bind heterologous IgG everywhere except to the tip (comprising approximately 10% of one fiber polypeptide) of the long tail fibers; (iii) cross-reacting OmpA-specific phages may bind heterologous IgG only to the tip of these fibers: (iv) OmpA-specific phages not cross-reacting at the tip of the tail fibers use different receptor sites on the protein. Absence of cross-reactivity appears to reflect high degrees of dissimilarity. A DNA probe consisting of genes encoding the two most distal tail fiber proteins of T4 detected homologies only in DNA from phages serologically cross-reacting at this fiber. Even under conditions of low stringency, allowing the formation of stable hybrids with almost 30% base mismatch, no such homologies could be found in serologically unrelated phages. Thus, in the collection of phages examined, there are sets of very similar and very dissimilar tail fiber genes and even of such gene segments.

Dynamics of non-linear biochemical systems and the evolutionary significance of time hierarchy.

Several non-linear reaction networks are analyzed in order to study the influence of time hierarchy on the dynamics of biochemical systems. The analysis is based on the assumption that the separation of the time constants within metabolic systems is a direct consequence of strong differences of enzyme concentrations. Therefore, as variable system parameters, only the enzyme concentrations are considered. By investigation of the stationary states of various three-component models with feedback-activation bifurcation diagrams within a two-dimensional parameter space, the enzyme simplex, are constructed. The diagrams contain the information about the number of stationary states, their stability properties as well as the type of motion expected at different parameter combinations. The results support the hypothesis that systems with separated time constants generally show a simple dynamic behaviour. Complex motions can be expected mainly for systems without time hierarchy, which are characterized by parameters located within the centre of the enzyme simplex. Quantitative measures for the time hierarchy and the complexity of the dynamics are derived. It is supposed that the separation of time constants is a main feature of the evolution of biochemical systems. The hypothesis is further supported by the consideration of the time hierarchy of the glycolytic pathway.

Cloning and expression in Escherichia coli K-12 of the genes for major outer membrane protein OmpA from Shigella dysenteriae, Enterobacter aerogenes, and Serratia marcescens.

The outer membranes of many gram-negative bacteria contain a major heat-modifiable protein which shows serological cross-reactivity with the OmpA protein of Escherichia coli K-12. Using the cloned gene for the E. coli K12 protein as a DNA-DNA hybridization probe, we were able to identify the corresponding genes from Shigella dysenteriae. Enterobacter aerogenes, and Serratia marcescens. These were cloned in a phage lambda vector, and their expression in E. coli K-12 was studied. All three OmpA proteins were fully produced and correctly exported to the outer membrane. In several cases, complete or partial restoration of known function of the E. coli K-12 protein was observed.

Mutants (ompA) affecting a major outer membrane protein of Escherichia coli K12.

Seventy independent mutants have been analyzed affecting a major protein, polypeptide II, of the outer cell envelope membrane from Escherichia coli K12. They were classified as nonsense mutants of the amber type (20%), mutants most likely of the missense type possessing the protein at normal concentrations (9%), and mutants either missing the protein or harboring it at much reduced concentrations for unknown reasons (71%). Forty of the mutants were analyzed genetically and all were found to map at or near ompA, the structural gene for protein II. Two-dimensional electrophoretic analyses of envelopes from such mutants revealed an unusual heterogeneity of the protein which on such patterns appeared as at least 12 well separated spots, and the majority of these is due to artifacts of the method but apparently specific for this protein. In no case was a polypeptide fragment found in envelopes from the nonsense mutants. The results are discussed regarding two different phages which use the protein as a receptor and concerning the biosynthetic incorporation of the protein into the outer membrane.

Cell envelope and shape of Escherichia coli: multiple mutants missing the outer membrane lipoprotein and other major outer membrane proteins.

Starting with an Escherichia coli strain missing the outer membrane lipoprotein, multiple mutants were constructed than in addition to this defect miss the outer membrane proteins II, Ia and Ib, or Ia, Ib, and II. In contrast to all single mutants or strains missing the lipoprotein and polypeptides Ia and Ib, drastic influences on the integrity of the outer membrane and cell morphology were observed in mutants without lipoprotein and protein II. Such strains exhibited spherical morphology. They required increased concentrations of electrolytes for optimal growth, and Mg2+ or Ca2+ were the most efficient. These mutants were sensitive to hydrophobic antibiotics and detergents. Electron microscopy revealed abundant blebbing of the outer membrane, and it could clearly be seen that the murein layer was no longer associated with the outer membrane.

Mutational change of membrane architecture. Mutants of Escherichia coli K12 missing major proteins of the outer cell envelope membrane.

Mutants of Escherichia coli have been analyzed which miss two of the major proteins of the outer cell envelope membrane. The two proteins I and II, normally are present at high concentrations (about 10(5) copies per cell). In such mutants, as compared with wild type, the phospholipid-to-protein ratio in the outer membrane has increased by a factor of 2.3 causing a considerable difference in density between wild type and mutant membranes. The concentrations of two other major components of the outer membrane, lipopolysaccharide and Braun's lipoprotein, did not change. The protein-deficient mutants do not exhibit gross functional defects in vitro. An increased sensitivity to EDTA and a slight such increase to dodecyl sulfate (but not to deoxycholate or Triton X-100) was observed, loss of so-called periplasmic enzymes was not found, and other differences to wild type are marginal. The mutants can grow with normal morphology. It is not possible, however, to prepare "ghosts" (particles of size and shape of the cell without murein, surrounded by a derivative of the outer membrane, and possessing the major proteins of this membrane) from them. This fact confirms our earlier suggestion that the proteins in question are required for the shape maintenance phenomenon in ghosts, and the mutants reject the speculation that these proteins are involved in the expression of the genetic information specifying cellular shape. Freeze-fracturing showed that in mutant cells, and in sharp contrast to wild type, the far predominant fracture plane is within the outer membrane. The concentration of the well known densely packed particles at the outer, concave leaflet of this fracture plane is greatly reduced. It was not possible, however, to clearly establish that one or the other protein is part of these particles because these ultrastructural differences were not apparent in mutants missing either one of the proteins only. The biochemical and ultrastructural data allow the conclusion that the loss of two major proteins and the concomitant increase of phospholipid concentration has changed the architecture of the outer membrane from a highly oriented structure, with a large fraction of protein-protein interaction, to one predominantly exhibiting planar lipid bilayer characteristics. E. coli thus can assemble rather different outer membranes, a fact excluding that outer membrane formation constitutes a highly ordered or strictly sequential assembly-line process.