Biochemical Characterization of Chlorophyll-Free Mitochondria From Pea Leaves

Abstract

Mitochondria from pea leaves were purified by centrifugation on a self-generated Percoll gradient which contained a linear gradient of polyvinylpyrrolidone-25 (0-10010, w/v). The chlorophyll content of the purified mitochondria was less than 1 pg per mg protein. All substrates were rapidly oxidized by these mitochondria, the rate of glycine oxidation being between 200 and 300 nmol 0, min-' mg-I protein, depending on the age of the leaves used. These rates did not vary significantly over a period of 20 h, provided NAD+ was supplied exogenously, when the mitochondria were stored on ice. Respiratory control, ADP/O ratios and outer membrane integrity (always more than 95010) were also maintained during storage. The phospholipid composition of the membranes from the leaf mitochondria was virtually identical to that of mitochondria from non-photosynthetic tissues although their lipid to protein ratio was slightly lower. The polypeptide pattern of the membranes from green leaf mitochondria and those from etiolated leaves and hypocotyls were also similar, but marked differences were observed between the matrix proteins from the different tissues. In particular, intensely stained bands at 94, 51,41 and 15.5 kDa which were present in the matrix of green leaf mitochondria were missing or present in much smaller quantities in the non-photosynthetic tissues. This difference was correlated with the ability of the mitochondria to oxidize glycine, suggesting that the four polypeptides may be associated with the glycine decarboxylase complex.

Developmental Physiology of Cluster-Root Carboxylate Synthesis and Exudation in Harsh Hakea. Expression of Phosphoenolpyruvate Carboxylase and the Alternative Oxidase

“…Cluster roots were excised from root systems and kept on ice until extraction (,1 h). Mitochondria were isolated from cluster roots, using a self-forming Percoll gradient, modified after Day et al (1985). Briefly, in a cold room (4°C) cluster roots were chopped into small pieces and then homogenized in grinding medium (0.3 M Suc, 25 mM tetrasodium pyrophosphate, 2 mM EDTA, 10 mM KH 2 PO 4 , and 1% [w/v] polyvinylpyrrolidone [PVP]-40) with a mortar and pestle.…”

Mannose-6-Phosphate Reductase, a Key Enzyme in Photoassimilate Partitioning, Is Abundant and Located in the Cytosol of Photosynthetically Active Cells of Celery (Apium graveolens L.) Source Leaves

“…The proteins that cross-reacted had molecular masses within the published range and were as follows: GD p subunit (Fig. ID) measured 95.9 kD, published range 94 to 98 kD (Day et al, 1985;Walker and Oliver, 1986); GO ( . Rubisco holoenzyme antibodies gave a strong signal with the large subunit, which measured 51.2 kD and has a published range of 51 to 58 kD (Jensen and Bahr, 1977) but only a weak signal with the small subunit, which meas- Lanes C through C, Total leaf protein (33 jjg) electroblotted to Immobilon P membranes and probed with antisera at the dilutions given below.…”

Specificity of the Organic Acid Activation of Alternative Oxidase in Plant Mitochondria

“…cv Stevens) were grown under glasshouse conditions in trays of vermiculite, and after 14 d the cotyledons were harvested. Mitochondria were isolated as described by Day et al (1985). Tubers of white sweet potato (lpomoea batatas L. var Leucorrhiza) were purchased from local markets.…”