Osmotic rupture

C. Osmotically Ruptured Intact Plastids (Ferricyanide Reduction)... 

The outer membranes of mitochondria can be removed from the inner membranes by osmotic rupture.13 Analyses on separated membrane fractions show that the outer membrane is less dense (density — 1.1 g / cm3) than the inner (density 1.2 g / cm3). It is highly permeable to most substances of molecular mass 10 kDa or less because of the presence of pores of 2 nm diameter. These are formed by mitochondrial porins,14-17 which are similar to the outer membrane porins of gram-negative bacteria (Fig. 8-20). The ratio of phospholipid to protein ( 0.82 on a weight basis) is much higher than in the inner membrane. Extraction of the phospholipids by acetone destroys the membrane. Of the lipids present, there is a low content of cardiolipin, a high content of phosphatidylinositol and cholesterol, and no ubiquinone. 

Since isolated thylakoids are less sensitive to hypnotic stress than intact cells, osmotic rupture does not occur to any significant extent during thawing of frozen thylakoids. 

Incubation of synaptosome preparations with labelled choline and isolation of the different constituents of the synaptosome by hypo-osmotic rupture and density gradient centrifugation showed labelling of the cytoplasmic ACh but very little labelling of the vesicular ACh (< 2%) (Marchbanks, 1969). Incubation of synapto-somes with labelled ACh showed that the cytoplasmic ACh but not the vesicular ACh was readily exchanged with external ACh. 

Enzymatic lysis CeU waU digestion and osmotic rupture Feasible... 

Papulacandin B, an antibiotic produced by the deuteromycete Papularia sphaerosperma, inhibits the growth of yeasts, but has no effect on other fungi, bacteria, or protozoa.The mode of action involves the inhibition of synthesis of alkali-insoluble jS-D-glucan of Saccharomyces cerevisiae and Candida albicans in sphaeroplasts and causes lysis of cells by osmotic rupture.Echinocandin B and a structurally related antibiotic aculeacin A also inhibit j3-D-glucan synthesis in yeast, possibly in a similar manner.Papulacandin B has been shown to inhibit the transfer of D-glucose from UDP-D-glucose, but not D-mannose from GDP-D-mannose, into dolichyl phosphate. 

Gasexchange of attached leaves of Phaseolus vulgavis and 820 nm absorbance changes, monitoring the redox state of P700 were measured simultaneously as in [4]. To supress photorespiration O2 was reduced to 2% when CO2 was low and net e"-transport from H2O to PGA) was calculated from CO2-uptake. Intact chloroplasts were isolated from Spinacia oleracea. For determination of the activation state of enzymes their activity was measured immediately after osmotic rupture of chloroplasts as in [5]. 

Electron microscopy of M. shows a characteristic internal structure. There are 2 concentric membranes, each 5-7 nm thick (see Biomembranes). Between the outer and inner membranes lies the intermembrane space (also called external matrix or outer mitochondrial space) (Figs. 1 2). These 2 membranes have different submicroscopic structures, their biogenesis is different, and they are functionally distinct. The outer membrane can be removed by osmotic rupture. 

Chloroplast envelope. Highly purified chloroplasts were isolated from leaves of wheat or spinach and, after osmotic rupture, subjected to density gradient centrifugation to isolate an envelope fraction [7]. This fraction was incubated with UDP[ H]galactose to label monogalactosyldiacylglycerol [7]. 

Enzyme activities were determined using established methods from the literature and were optimized to give the maximum rate. The recovery of enzyme activity in the washed plastids was determined as a percentage of that in the initial homogenate. The latency of enzyme activity was determined by assaying the enzyme activities using intact plastids under iso-osmotic conditions, and using plastids that had been osmotically ruptured. 

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