Acyl-CoA Synthetase Is Located in the Outer Membrane and Acyl-CoA Thioesterase in the Inner Membrane of Pea Chloroplast Envelopes

Abstract

Both acyl-CoA synthetase and acyl-CoA thioesterase activities are present in chloroplast envelope membranes. The functions of these enzymes in lipid metabolism remains unresolved, although the synthetase has been proposed to be involved in either plastid galactolipid synthesis or the export of plastid-synthesized fatty acids to the cytoplasm. We have examined the locations of both enzymes within the two envelope membranes of pea (Pisum sativum var Laxton's Progress No. 9) chloroplasts. Inner and outer envelope membranes were purified from unfractionated envelope preparations by linear density sucrose gradient centrifugation. Acyl-CoA synthetase was located in the outer envelope membrane while acyl-CoA thioesterase was located in the inner envelope membrane. Thus, it seems unlikely that the synthetase is directly involved in galactolipid assembly. Instead, its localization supports the hypothesis that it functions in the transport of plastid-synthesized fatty acids to the endoplasmic reticulum.

Testing Models of Fatty Acid Transfer and Lipid Synthesis in Spinach Leaf Using in Vivo Oxygen-18 Labeling

“…Thomas and co-workers have suggested that fatty acyl groups may be exported from the plastid as acyl carnitines (Thomas et al, 1982(Thomas et al, , 1983McLaren et al, 1985). In addition, an acyl-CoA thioesterase was observed with chloroplast envelopes (Joyard and Stumpf, 1980) and was later more specifically localized to the inner envelope (Andrews and Keegstra, 1983;Block et al, 1983). The in vivo function of this thioesterase has remained unclear.…”

“…Subsequent experiments showed that ACP-dependent fatty acid synthesis was entirely a chloroplast function (Ohlrogge et al, 1979), implying that the acyl-ACP thioesterase was a stromal enzyme activity. Acyl-CoA synthetase was demonstrated as an activity of the chloroplast envelope (Roughan and Slack, 1977;Joyard and Stumpf, 1981), and later more specifically of the outer envelope (Andrews and Keegstra, 1983;Block et al, 1983). The assumption is that free fatty acids are the intermediates, although the mechanism by which such intermediates move is unknown.…”

Expression of the Yeast Δ-9 Fatty Acid Desaturase in Nicotiana tabacum

“…Most polyunsaturated 18-carbon plant fatty acids, however, appear to be formed in the cytosol by ER-bound enzymes. Fatty acids that are transported out of the chloroplast are thought to be converted to CoA esters and subsequently incorporated into glycerolipids by the eukaryotic route (2,6,18,26). These lipids characteristically contain 18-carbon fatty acids in the sn-2 position.…”

Acyl Carrier Protein (ACP) Import into Chloroplasts

“…Hence, it has been proposed that this increase is due to the stimulation of acylCoA synthetase, a thermolysin-resistant enzyme (Dome et al, 1982) in the outer membrane of the chloroplast (Andrews and Keegstra, 1983;Block et al, 1983). In a CoA-and ATPdependent manner, acyl-CoA synthetase mediates the transport of free fatty acids from the chloroplast into the cytosol after their release by an inner membrane acyl-ACP thioesterase (Andrews and Keegstra, 1983;Block et al, 1983).…”

“…Earlier studies on the synthesis of fatty acids in plants suggested that CoA was not translocated into the chloroplast (Roughan et al, 1979;Weaire and chondria of plants (Neuburger et al, 1984) and animals (Tahiliani, 1989;Tahiliani and Neely, 1987) has been shown. Exogenously added CoA, on the other hand, does increase chloroplast synthesis of fatty acids and long-chain acyl-CoAs (Roughan et al, 1979;Sanchez and Mancha, 1981 ;Gardiner et al, 1984), perhaps through stimulating acyl-CoA synthetase, located in the outer envelope (Andrews and Keegstra, 1983;Block et al, 1983). In the present study we have examined how CoA and Ado(3',5')PZ affect the synthesis of holoACP during chloroplast import.…”