Regulation of rat liver acetyl-CoA carboxylase. Stimulation of phosphorylation and subsequent inactivation of liver acetyl-CoA carboxylase by cyclic 3':5'-monophosphate and effect on the structure of the enzyme.

The effects of citrate and cyclic AMP on the rate and degree of phosphorylation and inactivation of rat liver acetyl-CoA carboxylase were examined. High citrate concentrations (10 to 20 mM), which are generally used to stabilize and activate the enzyme, inhibit phosphorylation and inactivation of carboxylase. At lower concentrations of citrate, the rate and degree of phosphorylation are increased. Furthermore, phosphorylation and enzyme inactivation are affected by cyclic AMP under these conditions. At high citrate concentrations, cyclic AMP has little or no effect on inactivation and phosphorylation of acetyl-CoA carboxylase. Phosphorlation and inactivation of carboxylase is accompanied by depolymerization of the polymeric form of the enzyme into intermediate and protomeric forms. Depolymerization of carboxylase requires the transfer of the gamma-phosphate group from ATP to carboxylase. Inactivation occurs in the absence of CO2, which indicates that phosphorylation of the enzyme is the cause of inactivation and depolymerization, i.e. carboxylation of the enzyme is not responsible for inactivation of the enzyme.     

Specific adhesion of rat hepatocytes to beta-galactosides linked to polyacrylamide gels

Rat hepatocytes, isolated by a collagenase perfusion technique, specifically bind to polyacrylamide gel containing covalently immobilized 6-aminohexyl beta-D-galactopyranosyl groups. Less than 5% of these cells bind to polyacrylamide or to gels with the following covalently linked ligands: 6-aminohexanol, or the 6-aminohexyl D-pyranosides of alpha-mannose, beta-glucose, beta-2-acetamido-2-deoxyglucose, beta-cellobiose, beta-maltose, or beta-melibiose. Cell binding to beta-D-galactoside gels occurs after a lag period at 37 degrees and 65 to 100% (depending on the cell preparation) of the cells adhere. The duration of the lag period is inversely related to the beta-D-galactoside content of the gel but preincubation of the cells at 37 degrees reduces the lag period. Cell-gel binding is a threshold phenomenon. Adhesion of cells to gels does not occur when the glycoside concentration is less than about 900 nmol per cm2 x 0.25 mm thick gel piece. Above this critical concentration, cell-gel binding occurs and becomes maximal when the concentration is increased by only 20%. If these in vitro results apply to cellular interactions in vivo, they suggest that slight changes in the levels of cell surface or extracellular matrix carbohydrates may profoundly influence the behavior of neighboring cells.     

Effects of orchinol and related phenanthrenes on the enzymic degradation of indole-3-acetic acid

Orchinol, hircinol, loroglossol and certain related phenanthrenes inhibited horseradish peroxidase-catalysed IAA degradation to a varied degree. Among     

Isolation and cultivation of spirochetes and other spiral-shaped bacteria associated with the cecal mucosa of rats and mice.

A method for the culture of spiral-shaped bacteria associated with the intestinal mucosa of rodents is described. The appearance in culture of a spiral organism from rat ceca and a spirochete from mouse ceca is illustrated; these organisms are morphologically similar to the major inhabitants of the cecal mucosa in each animal species.     

Kinetic studies of enzymatic hydrolysis of insoluble cellulose: analysis of the initial rates

A study was conducted on the kinetics of enzymatic hydrolysis of pure insoluble cellulose using unpurified culture filtrate Trichoderma reesei, with the emphasis on the initial reaction period. The initial hydrolysis rate and extent of enzyme (soluble protein)adsorption, either apparent or initial, were evaluated under various experimental conditions. It has been found that the various mass-transfer steps do not control the overall hydrolysis rate and that the hydrolysis rate is mainly controlled by the surface reaction step promoted by the adsorbed enzyme. It has also been found that the initial hydrolysis rate strongly depends on the initial extent of soluble protein adsorption and the effectiveness of the adsorbed soluble protein to promote the hydrolysis. The initial extent of soluble protein adsorption, in turn, is related to the initial cellulose concentration, enzyme concentration, and specific surface area of cellulose, whereas the effectiveness of the initially adsorbed soluble protein to promote the derived to interrelate these parameters without resorting to the Michaelis-Menten kinetics. The present result appear to imply that the role of enzyme-substrate complex formation should not be ignored in deriving a mechanistic kinetic model for enzymatic hydrolysis of cellulose.     

Annular and non-annular binding sites on the (Ca2+ + Mg2+)-ATPase

Quenching of the fluorescence of the $\text{(}\text{Ca}{}^{\mathrm{2+}}\text{+}\text{Mg}{}^{\mathrm{2+}}\text{)-}\text{ATPase}$ purified from muscle sarcoplasmic reticulum can be used to measure relative binding constants of hydrophobic compounds to the phospholipid-protein interface. We show that the binding constant for cholesterol is considerably less than that for phosphatidylcholine, so that cholesterol is effectively excluded from the phospholipid annulus around the ATPase. However, dibromocholestan-3β-ol causes quenching of the fluorescence of the ATPase, and so has access to other, non-annular sites. We suggest that these non-annular sites could be at protein/protein interfaces in ATPase oligomers. Oleic acid can bind at the phospholipid/protein interface, although its binding constant is less than that for a phosphatidylcholine, and it can also bind at the postulated non-annular sites. The effects of these compounds on the activity of the ATPase depend on the structure of the phospholipid present in the systems.