Radiation inactivation analysis of H⁺‐pyrophosphatase from submitochondrial particles of etiolated mung bean seedlings

Abstract

Radiation inactivation analysis was employed to determine the functional masses of enzymatic activity and proton translocation of H + -pyrophosphatase from submitochondrial particles of etiolated mung bean seedlings. The activities of H + -pyrophosphatase decayed as a simple exponential function with respect to radiation dosage. D 37 values of 6.9 þ 0.3 and 7.5 þ 0.5 Mrad were obtained for pyrophosphate hydrolysis and its associated proton translocation, yielding molecular masses of 170 þ 7 and 156 þ 11 kDa, respectively. In the presence of valinomycin and 50 mM KCl, the functional size of H + -pyrophosphatase of tonoplast was decreased, while that of submitochondrial particles remained the same, indicating that they are two distinct types of proton pump using PP i as their energy source.z 2000 Federation of European Biochemical Societies.

Radiation inactivation analysis of thylakoid protein kinase systems in light and in darkness

“…Irradiation was performed with a 60 Co irradiator (approximately 1.11×10 6 GBq) and the dose rate was 7.00×10 -3 Mrad s -1 determined as previously described (Pan et al 1987, Wang et al 1988, Ma et al 1993, Tzeng et al 1996, Jiang et al 2000, Wu et al 2004. Internal standardization of radiation exposure was carried out with G 6 PDH.…”

“…Radiation inactivation analysis has been used effectively to estimate the functional mass and thus to elucidate the structure-function relationship of many biological molecules and systems including ATP synthase (Wang et al 1988, Ma et al 1993, HIV-1 reverse transcriptase (Sluis-Cremer et al 2003), photosynthetic electron transport chain (Pan et al 1987), progesterone receptor (Wu et al 2004), pyrophosphatase (Jiang et al 2000), and vertebrate hyaluronan synthase (Pummill et al 2001). In this work, we report the functional masses of thylakoid protein kinase system(s) (TPKXs) determined by radiation inactivation technique.…”

Proton pumping inorganic pyrophosphatase of endoplasmic reticulum-enriched vesicles from etiolated mung bean seedlings

“…Possible existence of a pyrophosphatase on plasma membranes from Ricinus communis seedlings (Long et al, 1995), cauliflower inflorescence cells (Ratajczak et al, 1999), and tobacco culture cells (Sikora et al, 1998) was suggested. In addition, several groups successfully observed a proton pumping activity of submitochondrial particles (Vianello et al, 1991;Jiang et al, 2000). A pyrophosphatase was also purified from thylakoid membrane of spinach (Jiang et al, 1997).…”

“…The molecular mass of their H + -PPase was markedly smaller than that from vacuole, indicating a possible new type of the H + -PPase on mitochondrial membranes (Zancani et al, 1995). However, radiation inactivation analysis demonstrated that submitochondrial particles from etiolated mung bean seedlings contained a H + -PPase with a functional size of 170 kDa (Jiang et al, 2000). Furthermore, it was revealed that the PPase purified from spinach thylakoid membrane lacked of any proton pumping activity (Jiang et al, 1997).…”

Pyrophosphate import and synthesis by plant mitochondria

“…1999). On the basis of radiation‐inactivation analysis, however, molecular masses of 170 and 156 kDa were, respectively, obtained for PPi hydrolysis and its associated proton translocation catalysed by the H + ‐PPiase of mung bean mitochondria (Jiang et al. 2000).…”