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S Speiser

Publications and source records attributed to S Speiser.

6 recordsLinked to original sources
PubMed1983

ATP stimulates proteolysis in reticulocyte extracts by repressing an endogenous protease inhibitor.

An endogenous inhibitor of the reticulocyte ATP-dependent proteolytic system has been purified partially by ammonium sulfate precipitation from rabbit reticulocyte and erythrocyte extracts. Inhibitor-free protease rapidly degrades 21-40% of the substrate [14C]methyl-alpha-casein per hour, resembling ATP-dependent activity in reticulocyte extracts. This proteolytic activity is not stimulated by ATP and does not respond to ubiquitin. Adding back the inhibitory fraction to reticulocyte inhibitor-free protease results in a significant decrease (65-75%) in proteolysis, both in the presence and absence of ATP. In contrast, inhibition is repressed when both ATP and the ubiquitin-containing fraction are present, resulting in an 80-350% stimulation of proteolysis by these components. These results suggest that ATP, in the presence of ubiquitin, may act in releasing the protease(s) from its endogenous inhibitor. Erythrocyte extracts, unlike reticulocyte extracts, exhibit low levels of ATP-dependent proteolytic activity. However, ion-exchange chromatography reveals that erythrocytes contain levels of proteolytic activity that are comparable to the reticulocyte's inhibitor-free protease. Addition of ubiquitin and inhibitor to erythrocyte protease results in a highly ATP-dependent activity that resembles levels of ATP-dependence (3- to 4-fold) seen in reticulocyte extracts. Thus, the proteolytic and inhibitory components of the ATP-dependent proteolytic system appear to be retained with reticulocyte maturation. However, some other component(s) of the system are lost or modified with maturation so that the protease remains inactive.

Adenosine Triphosphate↗
PubMed1983

Regulation of proteolysis in erythroid cells.

Reticulocytes contain non-lysosomal ATP dependent proteolytic activity which appears important in degrading abnormal proteins as well as certain organelles. ATP acts to repress an endogenous protease inhibitor activity by a process requiring the polypeptide ubiquitin. Free epsilon amino groups on substrates are important for recognition by the full ATP-dependent system but do not appear necessary for hydrolysis per se suggesting that ubiquitin-substrate conjugates repress the inhibitor. ATP-dependent proteolysis declines markedly with reticulocyte maturation and decreases further to negligible levels with aging of erythrocytes. With cell maturation the inhibitor and protease(s) remain but the ubiquitin-containing fraction is less effective in repressing the inhibitor. Finally, additional control mechanisms selective for certain proteins may exist.

Adenosine Triphosphate↗
PubMed1981

ATP-dependent proteolysis in erythroid and muscle cells.

Reticulocytes contain a soluble, non-lysosomal proteolytic activity which appears responsible for the energy-dependent degradation of several proteins within these cells. This activity requires Mg++-ATP, has a pH-optimum of 7.8, and is inhibited by hemin and sulfhydryl reagents such as iodoacetamide and N-ethylmaleimide. With reticulocyte maturation ATP-dependent activity disappears although protease not requiring ATP remains and even increases. Poly-L-lysine was found to specifically inhibit ATP-dependent activity although it is a poor substrate for this system. Various protease fractions with reduced ATP-dependence and erythrocyte activity not requiring ATP showed decreased sensitivity to polylysine. The effect of polylysine was found to increase with polymer length and may relate to the apparent selectivity of ATP-dependent degradation toward large substrates. In addition, reductive methylation of amino groups on alpha-casein selectively inhibited ATP-dependent breakdown of this protein. Such results suggest that ATP may act at a step which proceeds proteolysis and that substrate amino groups play a role in recognition by the ATP-dependent system. However, such experiments do not yet exclude other mechanisms of action for polylysine's effect. Other experiments have confirmed that reticulocyte activity may involve a non-proteolytic factor which stimulates a protease-containing fraction from these cells. An analogous ATP-dependent activity has been described in muscle suggesting a general function for this degradative activity. Future studies should lead to more insight into the role of ATP and the importance of this system in protein catabolism.

Adenosine Triphosphate↗