Detergent/protein ratio

In several tissues, like gastric mucosa [6,8,46.47], pancreas [19,20], submandibular gland [22] and intestinal mucosa [36], the anion-sensitive ATPase can be solubilized by Triton X-100, a detergent/protein ratio of 3 1 being most effective [6,8,47]. The solubihzed enzyme from gastric mucosa [46] and pancreas [20] could be partially purified by gel filtration and sucrose density gradient centrifugation. The properties of the solubilized enzyme do not differ very much from those of the particulate enzyme. 

The Arl2 GAP activity is solubilized by bringing the crude mitochondrial membranes to a final concentration of a 2% CHAPS at a detergent protein ratio of 2.5 detergentil total protein (w/w) and incubating for 30 min on ice. The sample is then spun at 100,000g for 1 h and the supernatant is recovered and stored at 4°. 

Analysis of the relative areas under the stable boundaries indicated that the composition of the complexes 1 and 2 was constant and that the distribution between the components was a linear function of the mixing ratio. The first complex, AD , containing 0.22 g. SDS per g. protein (or the equivalent of half the cationic groups of serum albumin) corresponds in composition to the precipitate at the minimum detei ent/ protein ratio requisite for complete precipitation. The second complex, AD2n, contains just twice as much SDS (the equivalent of the total acidbinding capacity) and corresponds in composition to the precipitate obtained at the maximum detergent/protein ratio yielding complete... 

Solubilize desaturases with 1 % CHAPS at a detergent-membrane protein ratio of equal to one, 5°C for 30 min with gentle stirring. 

Method Structure class Protein Size, kDa ofTM a (13) Detergent Detergent protein molar ratio Temp C PDB ID Ref... 

The cytochrome b559 content was found to be 1 per 2 pheophytin a (19) and subsequent analyses have shown the same ratio in larger PS2 preparations (20). The fatty acid content has been analysed and indicates that there is always less than 1.0 mole dlacyl glycerolipid 2 pheophytin a. This low level probably arises from non-specific interaction with the detergent-protein complex. 

Table 10.4 Detergent/protein (w/v) ratios for 90% maximal solubilization of membrane protein...

Fiq. 2. Precipitation, of horse serum albumin by sodium dodecyl sulfate as a function of detergent/protein weight ratio at pH 4.5 in 0.1 N sodium acetate buffer. Adapted from Putnam and Neurath (114). 

Viscosity measurements by Neurath and Putnam (101) have verified the existence of interaction in protein-detergent mixtures and have revealed a unique dependence of the viscosity increment on detergent/ protein weight ratio. This work showed that both cationic and anionic detergents produce a large increase in the relative viscosity of serum... 

To display total chlorophyll-protein complexes, corn thylakoids were washed with cold 2 mM Tris-maleate, pH 8.0, solubilized with a mixture of 2% octyl glucoside and 0.5% SDS to give detergent/Chi ratios of 40 and 10 respectively and electrophoresed on 10% polyacrylamide gels in the presence of 0.1% SDS (Camm, Green, 1980). 

The 26 kDa protein synthesised by salt-adapted tobacco cells has been further characterised (Singh et al., 1987a). The protein makes up approximately 12% of the total cellular protein and has been resolved into two forms. These two forms have been designated osmotin 1 and osmotin II and occur in a 2 3 ratio. The forms are distinct with osmotin I soluble in an aqueous phase and osmotin II soluble in detergent. The proteins accumulate as inclusion bodies in the vacuole and are only sparsely distributed in the cytoplasm. 

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