Lemire B D, Robinson J J, Bradley R D, Scraba D G, Weiner J H
J Bacteriol. 1983 Jul;155(1):391-7. doi: 10.1128/jb.155.1.391-397.1983.
The terminal electron transfer enzyme fumarate reductase has been shown to be composed of a membrane-extrinsic catalytic dimer of 69- and 27-kilodalton (kd) subunits and a membrane-intrinsic anchor portion of 15- and 13-kd subunits. We prepared inverted membrane vesicles from a strain carrying the frd operon on a multicopy plasmid. When grown anaerobically on fumarate-containing medium, the membranes of this strain are highly enriched in fumarate reductase. When negatively stained preparations of these vesicles were examined with an electron microscope, they appeared to be covered with knob-like structures about 4 nm in diameter attached to the membrane by short stalks. Treatment of the membranes with chymotrypsin destroyed the 69-kd subunit, leaving the 27-, 15-, and 13-kd subunits bound to the membrane; these membranes appeared to retain remnants of the structure. Treatment of the membranes with 6 M urea removed the 69- and 27-kd subunits, leaving the anchor polypeptides intact. These vesicles appeared smooth and structureless. A functional four-subunit enzyme and the knob-like structure could be reconstituted by the addition of soluble catalytic subunits to the urea-stripped membranes. In addition to the vesicular structures, we observed unusual tubular structures which were covered with a helical array of fumarate reductase knobs.
末端电子传递酶延胡索酸还原酶已被证明由一个由69千道尔顿（kd）和27千道尔顿亚基组成的膜外在催化二聚体以及一个由15千道尔顿和13千道尔顿亚基组成的膜内在锚定部分组成。我们从一个在多拷贝质粒上携带frd操纵子的菌株制备了反向膜泡。当在含延胡索酸的培养基上厌氧生长时，该菌株的膜中富含延胡索酸还原酶。当用电子显微镜检查这些膜泡的负染制剂时，它们似乎被直径约4纳米的旋钮状结构覆盖，这些结构通过短柄附着在膜上。用胰凝乳蛋白酶处理膜会破坏69千道尔顿的亚基，使27千道尔顿、15千道尔顿和13千道尔顿的亚基与膜结合；这些膜似乎保留了结构的残余部分。用6 M尿素处理膜会去除69千道尔顿和27千道尔顿的亚基，使锚定多肽保持完整。这些膜泡看起来光滑且无结构。通过向用尿素剥离的膜中添加可溶性催化亚基，可以重建一种功能性的四亚基酶和旋钮状结构。除了膜泡结构外，我们还观察到了不寻常的管状结构，这些结构被延胡索酸还原酶旋钮的螺旋阵列覆盖。
