The Chlamydomonas cell wall degrading enzyme, lysin, acts on two substrates within the framework of the wall

The Chlamydomonas cell wall is a multilayered, extracellular matrix containing 20-25 proteins and glycoproteins, many of which are highly enriched in hydroxyproline. 80-90% of the wall protein is located in a crystalline portion of the wall that is soluble in sarkosyl-urea solutions as well as in chaotropic salts. Although the wall has no cellulose it contains a noncrystalline, highly insoluble framework portion that is responsible for the integrity and overall shape of the wall. In the present report we show that the framework of the wall is composed of two components that are acted upon by lysin, a wall degrading enzyme released by mating gametes. One, which makes up the major portion of the framework, is insoluble upon boiling in SDS-PAGE sample buffer. Lysin treatment of this portion leads to its physical degradation and the concomitant appearance of several SDS-dithiothreitol-soluble polypeptides ranging in relative molecular mass from greater than 400,000 to less than 60,000. The second component is the flagellar collar. This hollow cylinder composed of striated fibers aligned in parallel array serves as the tunnel in the wall through which the flagella protrude. Our evidence indicates that the primary collar polypeptide is a 225,000-Mr molecule that itself has at least two functional domains. One domain, contained in a 185,000-Mr fragment, permits the self-association of the molecules to form the main body of the collar. The second part of the molecule anchors the collar to the wall framework via sarkosyl-urea-insensitive, SDS-dithiothreitol-sensitive linkages.     

The thermal induced helix--beta transition of poly(N epsilon-methyl-L-lysine) and poly(N delta-ethyl-L-ornithine) in aqueous solution

Uncharged poly(N^ε-methyl-L -lysine) (PMLL) and its isomer, poly(N^δ-ethyl-L -ornithine) (PELO), in alkaline solution (pH ca. 12) undergo a helix-to-β transition upon mild heating at 50°C or higher in a manner similar to that of poly(L -lysine) (PLL). The rate of conversion follows the order: PMLL < PELO < PLL. The helix can be regenerated upon cooling near zero degrees, for instance, after more than 12 hr at 2°C. At concentrations less than 0.02% the β form is intramolecular, but at higher concentrations both intra- and intermolecular β forms are generated. Poly(N^δ-methyl-L -ornithine) (PMLO), an isomer of PLL, behaves like poly(L -ornithine); uncharged PMLO in alkaline solution is partially helical and becomes disordered at elevated temperatures.     

Genetic and Physical Mapping of the Mcra (Rgla) and Mcrb (Rglb) Loci of Escherichia Coli K-12

We have genetically analyzed, cloned and physically mapped the modified cytosine-specific restriction determinants mcrA (rglA) and mcrB (rglB) of Escherichia coli K-12. The independently discovered Rgl and Mcr restriction systems are shown to be identical by three criteria: 1) mutants with the RglA- or RglB- phenotypes display the corresponding McrA- or McrB- phenotypes, and vice versa; 2) the gene(s) for RglA and McrA reside together at one locus, while gene(s) for RglB and McrB are coincident at a different locus; and 3) RglA+ and RglB+ recombinant clones complement for the corresponding Mcr-deficient lesions. The mcrA (rglA) gene(s) is on the excisable element e14, just clockwise of purB at 25 min. The mcrB (rglB) gene(s), at 99 min, is in a cluster of restriction functions that includes hsd and mrr, determinants of host-specific restriction (EcoK) and methyladenine-specific restriction respectively. Gene order is mcrB-hsdS-hsdM-hsdR-mrr-serB. Possible models for the acqusition of these restriction determinants by enteric bacteria are discussed.     

Enzyme catalyzed hydrolysis of inorganic pyrophosphate. Current view of problems in characterization of PP1-phosphohydrolytic activity associated with alkaline phosphatases (EC 3.1.3.1).

A survey of the hydrolytic activity of alkaline phosphatase (EC 3.1.3.1) reveals that PP1, like phosphomonoesters, can serve as substrate in vitro. This pp1-phosphohydrolytic activity can be distinguished from PP1-phosphohydrolytic activities of inorganic pyrophosphatases (EC 3.6.1.1) and glucose-6-phosphatase (EC 3.1.3.9) by several criteria. Discrimination among these hydrolytic enzymes is possible by their dependence on variation of pH and of magnesium to PP1 ratios in the assay solutions. The true substrates and modifiers are not simply PP1 and magnesium, but the equilibrium species in mixtures of these two. The physiological significance of each of the three enzymes is not predictable from their differential efficiency as catalysts of PP1-hydrolysis in vitro.