Composition and enzyme activities of Spiroplasma citri membranes

Abstract

Spiroplasma citri was cultured in three different media that supplied cholesterol and fatty acids from: (i) horse serum, (ii) pleuropneumonia-like organism (PPLO) serum fraction, or (iii) bovine serum albumin-fatty acid-cholesterol. The ability of PPLO serum fraction to support growth varied by lot number. Neither PPLO serum fraction nor the bovine serum albumin medium supported growth as well as the horse serum medium. Analysis of cholesterol, lipid phosphorus, and membrane protein showed the horse serum- and PPLO-grown cells to be indistinguishable, but the bovine serum albumin-grown cells were deficient in lipid phosphorus. The three cultures did not show markedly different fatty acid compositions, but, in all cases, the cultures preferentially incorporated palmitic acid and discriminated against linoleic acid. Cultures grown for different times from logarithmic growth through a degenerative phase showed relatively constant ratios of cholesterol/protein and lipid phosphorus/protein. Fatty acid composition was also relatively constant at the different stages. Adenosine triphosphatase and p-nitrophenyl phosphatase were mainly associated with the membrane, whereas reduced nicotinamide adenine dinucleotide oxidase was either readily removed or not associated with the membrane. The reduced nicotinamide adenine dinucleotide oxidase was inactivated at temperatures above 35 degrees C.

Comparison of the membrane composition of Spiroplasma citri and the corn stunt Spiroplasma

“…Table 2 shows the fatty acid analyses for the membrane lipids of the two species. They are quite similar and in agreement with the previous analyses of S. citri (7). The main features are that the spiroplasmas select palmitate from the growth medium and discriminate against laurate (12:0), myristate (14:0), and linoleate (18:2).…”

“…S. citri membranes contained slightly more cholesterol and less lipid phosphorus than did membranes of the corn stunt spiroplasma, but the ratio of lipid to protein was about the same in the two organisms. Note, however, that these analyses for S. citri (Cir 3B grown at 28°C) show more cholesterol and less lipid phosphorus than the previous analyses (C189 grown at 32°C) (7). Strains C189 and Cir 3B were grown simultaneously in the medium used for the comparison of Cir 3B and corn stunt spiroplasma, at 30°C.…”

“…Previously we analyzed the lipid composition and enzyme activities of S. citri membranes (3,7). We have now studied these properties in the corn stunt spiroplasma and here report a comparison of corn stunt spiroplasma and S. citri.…”

“…For each analysis of corn stunt spiroplasma, 12 tubes (total volume 340 ml) were pooled, and final pH's were 6.86 to 6.88. Membranes were harvested and washed as described previously (7). Protein, lipid phosphorus, cholesterol, fatty acid, and adenosine triphosphatase were assayed as described previously (7).…”

“…Membranes were harvested and washed as described previously (7). Protein, lipid phosphorus, cholesterol, fatty acid, and adenosine triphosphatase were assayed as described previously (7). NADH diaphorase was measured using 2,6-dichlorophenolindophenol as an electron acceptor (6).…”