Stroma proteins

Rodoni, S. et al.. Partial purification and characterization of red chlorophyll catabolite reductase, a stroma protein involved in chlorophyll breakdown. Plant Physiol, 115, 677, 1997. 

FIGURE 1. Thin section of part of an isolated chloroplast showing the internal thylakoid membrane system which consists of appressed grana lamellae (g) and non-appressed stroma lamellae (s) embedded in the stroma protein matrix and surrounded by a double membrane envelope (e). 

The amount of stroma proteins is less in fish muscles (3-5%) than it is In beef or rabbit muscles (15-18%). This may explain why raw fish fillets are acceptable in Japanese dishes, whereas beef, rabbit and pork are rarely served raw. According to Fennema et al. (9.), tenderness is primarily related to collagen content, while toughness and water-holding capacity are associated with the myofibrillar proteins. Many papers on cooked meat mention both tenderness and toughness, while those on cooked fish note the problems of toughness rather than tenderness. This also might be related to the difference in content of the stroma proteins. 

Rodoni S, Vicentini F, Schellenberg M, Matile P, Hortensteiner S (1997) Partial Purification and Characterization of Red Chlorophyll Catabolite Reductase, a Stroma Protein Involved in Chlorophyll Breakdown. Plant Physiol 115 677... 

The actomyosin content of fish muscle, some 70%, appears to be appreciably greater than that of mammals. The difference may well reside in the muscle as a whole rather than in the fiber content. If it is assumed that the stroma protein consists substantially of sarcolemma, connective tissue, and blood vessels, then the difference would be due to... 

The pellet was washed twice with 50 mM Ttis-HCl buffer, pH 8.0, containing 10 mM NaCl, to purify the membranes from stroma proteins. The thylakoid membranes were then homogenized with a small amount of washing buffer and sonicated in a cold ulttasonic bath for 30 min. For fiimre solubilization of the membrane proteins an equal volume of cold n-butanol (-20°Q was used. The phases were separated by centrifugation for 5 min at 1000 g and vratet phase contained the proteins was collected. The butanol extraction was repeated twice to obtain a lipid-free protein preparation. 

There is clearly no exclusive site of acylation of glycerol phosphate in the plant cell. Density gradient centrifugation of the 270-g supernatant from castor bean endosperm showed the acylation activity mainly in the ER fraction (ca. 90%) and the remainder in the mitochondria (Vick and Beevers, 1977). However, it is clear from the results cited above that acylation activity is present in the chloroplast envelope (Joyard and Douce, 1977) and in the stroma proteins from chloroplasts (Joyard and Douce, 1977). 

In their studies on the enzymatic properties of chloroplast fractions, Joyard and Douce (1977) found an active PA phosphatase associated with the chloroplast envelope. Under some conditions monoglyceride is produced when stroma proteins and envelope together are incubated with labeled i/z-glycerol-3-P, perhaps demonstrating the action of a monoacylgly-cerol-3-P (LPA) phosphatase. 

A titration curve was established by the ELISA technique which shows the proportionnality between absorbance at 405 nm and various quantities of pure acyltranferase in the 1 to 25 ng range. This technique allows a quantification of the acyltransferase in chloroplast fractions (Fig. 3). This very active protein represents less than 0.04% of the stroma proteins. 

Phytyl-PP is formed from GGPP by hydrogenation at the envelope membrane / / (Fig, 3). Another pathway is the stepwise hydrogenation of GG-chloro-phyllide a to form chlorophyll a at thylakoid membranes /17/ (Fig. 3), NADPH functions as electron donor in both reactions. A kinase which forms phytyl-PP from phytol plus ATP is localized in the stroma /6/. GGPP Itself is formed from IPP by a recombinated system of envelope or thylakoid membranes plus stroma protein /I8/,... 

Lipid release from immobilized chloroplast envelope required stroma and was stimulated by ATP, GTP and palmiloyl-CoA (Table 1). As with ER, other acyl-CoAs were stimulatory and NEM was without effect. GTPyS abolished both the GTP effect as well as a significant proportion of the stroma effects (Table 1). Thus, the stimulatory effect of GTP probably was connected with GTP hydrolysis interacting with stroma protein(s) rather than with a GTP-binding protein. Since stroma inhibits envelope to envelope vesicular transfer [6], lipid release from the envelope in the presence of stroma probably was not underestimated. The result supports previous findings of a metabolically regulated lipid transport between envelope and thylakoids [6]. 

A further complication of these reactions is that many nonhemoglobin proteins contain reactive groups and may also be modified to produce new, potentially toxic, contaminants. It has been difficult to produce a pure modified hemoglobin for toxicity studies because most processes start with relatively cmde stroma-free hemoglobin. 

The initial conversion of light into chemical energy takes place in the thylakoid membrane. Besides the chlorophylls and series of electron carriers, the thylakoid membrane also contains the enzyme adenosine triphosphate (ATP) synthase. The enzymes that are responsible for the actual fixation of C02 and the synthesis of carbohydrate reside in the stroma that surround the thylakoid membrane. The stroma also contains deoxyribonucleic acid (DNA), ribonucleic acid (RNA), and ribosomes that are essential for protein synthesis [37]. 

Chloroplasts (29-36) are the sites of photosynthesis and their ribosomes can carry out protein synthesis. Chloroplasts that contain chlorophylls and carotenoids, are disc shaped and 4-6 pm in diameter. These plastids are comprised of a ground substance (stroma) and are traversed by thylakoids (flattened membranous sacs). The thylakoids are stacked as grana. In addition, the chloroplasts of green algae and plants contain starch grains, small lipid oil droplets, and DNA. 

Like the Tom and Tim systems on mitochondrial outer and inner membranes, chloroplasts use the Toe and Tic systems on their outer and inner envelope membranes. Although there may not be a direct correspondence between both subunits, their functions for protein translocation appear quite similar. Thus, most of the sorting mechanisms within the envelope membranes are recognized as variations of the general sorting pathway to the stroma. 

The inner envelope membrane proteins have a cleavable N-terminal transit peptide, as well as some hydrophobic domain (s) in their mature portion. There are two possibilities on the role of this hydrophobic domain it may work as an N-terminal signal peptide after the translocation into the stroma and the subsequent cleavage of the transit peptide. Alternatively, it may work as a stop-transfer signal. One more important question is how the distinction is made between the outer membrane proteins, the inner membrane proteins, and the thylakoid membrane proteins. It is still an enigma. 

All thylakoidal proteins seem to be first translocated into the stroma through the previously mentioned general import pathway all of them have a cleavable N-terminal transit peptide. However, there are at least four different pathways into the thylakoid membrane (Robinson et al, 1998 Schnell, 1998). Most of them are reminiscent of the pathways of bacteria, described in Section II,B,1. It is not surprising because chloroplasts are most likely evolved from a prokaryotic endosymbioint, but there are certain differences. 

Two reasons could explain the failure of soluble SI SCF to induce long term growth of hematopoietic cells on stroma. One might be the production of inadequate levels of protein by SI stroma and by SESl and MMCE cells ectopically expressing SI SCF. The other could be an antagonistic function of the mutated SI SCF compared to the membrane-bound isoform. To test for these possibilities we added recombinant soluble SCF, that is produced as a cleavage product of mb SCF, to TEl feeder cocultures. 

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