The phosphoenolpyruvate-dependent fructose-specific phosphotransferase system in Rhodopseudomonas sphaeroides. Distribution of EIIFru over the membranes of phototrophically grown Rps. sphaeroides

The distribution of the fructose carrier over the membranes of Rhodopseudomonas sphaeroides was studied in cells grown under light saturation and light limitation. Three types of membranes were isolated after disruption of the cells in a French press. All three types were present in the cells grown either under the high or low light intensity, but they were present in different quantities. The cytoplasmic membrane could be separated from the photosynthetic membranes by Sephacryl S-1000 chromatography. The cytoplasmic membrane has the highest specific density and fructose carrier content and does not contain the light-harvesting pigments. The photosynthetic membranes could be resolved into two types by sucrose density gradient centrifugation. Type A predominates when cells are grown under light saturation, whereas type B, the chromatophores, is synthesized abundantly under light limitation. The properties of type A are in between the properties of the cytoplasmic membrane and the chromatophores. It has a slightly lower specific density and contains four times less fructose carrier than the cytoplasmic membrane, but contains half of the light-harvesting bacteriochlorophyll of the chromatophore membrane. The fructose carrier content in the type B membranes, the chromatophores, is very low.     

The electrochemical H+ gradient in the yeastRhodotorula glutinis

The electrochemical gradient of protons, , was estimated in the obligatory aerobic yeastRhodotorula glutinis in the pH₀ range from 3 to 8.5. The membrane potential, , was measured by steady-state distribution of the hydrophobic ions, tetraphenylphosphonium (TPP⁺) for negative above pH₀ 4.5, and thiocyanate (SCN^–) for positive below pH₀ 4.5. The chemical gradient of H⁺ was determined by measuring the chemical shift of intracellular P_i by³¹P-NMR at given pH₀ values. The values of pH_i increased almost linearly from 7.3 at pH₀ 3 to 7.8 at pH₀ 8.5. In the physiological pH₀ range from 3.5 to 6, was fairly constant at values between 17–18 KJ mol^–1, gradually decreasing at pH₀ above 6. In deenergized cells, the intracellular pH_i decreased to values as low as 6, regardless of whether the cell suspension was buffered at pH₀ 4.5 or 7.5. There was no membrane potential detectable in deenergized cells.     

Interspecies transduction of plasmids among Bacillus anthracis, B. cereus, and B. thuringiensis

Bacteriophage CP-51, a generalized transducing phage for Bacillus anthracis, B. cereus, and B. thuringiensis, mediates transduction of plasmid DNA. B. cereus GP7 harbors the 2.8-megadalton multicopy tetracycline resistance plasmid, pBC16. B. thuringiensis 4D11A carries pC194, the 1.8-megadalton multicopy chloramphenicol resistance plasmid. When phage CP-51 was propagated on these strains, it transferred the plasmid-encoded antibiotic resistances to the nonvirulent Weybridge (Sterne) strain of B. anthracis, to B. cereus 569, and to strains of several B. thuringiensis subspecies. The frequency of transfer was as high as 10(-5) transductants per PFU. Tetracycline-resistant and chloramphenicol-resistant transductants contained newly acquired plasmid DNA having the same molecular weight as that contained in the donor strain. Antibiotic-resistant transductants derived from any of the three species were effective donors of plasmids to recipients from all three species.     

Chromosomal mapping of Bacillus thuringiensis by transduction.

Three groups of linked markers were mapped in Bacillus thuringiensis 4042B by using two-, three-, and four-factor crosses mediated by the temperate bacteriophages TP-13 and TP-18. The order of markers was (trp-11, trp-2)-(leu-1, leu-2)-his-1-(lys-1, lys-2)-cys-1 in the first group; met-1-(argCl, argOl)-met-2-(pyr-1, pyrA2) in the second group; and met-3-pur-1-(nal-1, nal-2)-str-1-(pur-2, pur-4)-pur-3 in the third group. Electron microscopic measurements of head sizes suggested that the volume of the TP-13 phage head is seven times greater than that of the TP-18 phage head. The TP-18 genome was shown by DNA restriction analysis to have a molecular mass of 36 megadaltons. TP-13 was useful for scanning large segments of the B. thuringiensis chromosome, and TP-18 was effective for ordering markers too closely linked for simple resolution with TP-13.     

Vicinal dithiol-disulfide distribution in the Escherichia coli mannitol specific carrier enzyme IImtl

Escherichia coli mannitol specific EII in membrane vesicles can be inhibited by the action of the oxidizable substrate-reduced phenazine methosulfate (PMS) in a manner similar to E. coli enzyme IIGlc [Robillard, G. T., & Konings, W. (1981) Biochemistry 20, 5025-5032]. The fact that reduced PMS and various oxidizing agents protect the enzyme from inactivation by the sulfhydryl reagents N-ethylmaleimide and bromopyruvate suggests that the active form possesses a dithiol which can be protected by conversion to a disulfide. The sulfhydryl-disulfide distribution has been examined in purified EIImtl by labeling studies with N-[1-14C]ethylmaleimide ( [14C]NEM). EIImtl can be alkylated at three positions per peptide chain. When alkylation takes place in 8 M urea, only two positions are labeled. The third position becomes labeled in urea only after treatment with DTT, suggesting that the native enzyme is composed of two subunits linked by a disulfide bridge. The remaining two sulfhydryl groups per peptide chain appear to undergo changes in oxidation state as indicated by the following results. (1) Treatment of the active enzyme with NEM leads to complete inactivation and incorporation of 1 mol of [14C]NEM per peptide chain. Oxidizing agents protect the activity and prevent labeling presumably by forming a disulfide. (2) Phosphorylating the enzyme (one phosphoryl group per peptide chain) fully protects the activity, but 1 mol of NEM per peptide chain is still incorporated. Subsequent dephosphorylation by adding mannitol causes a second mole of [14C]NEM to be incorporated and results in complete inactivation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Diabetic Neuroarthropathy: Report of Four Cases

Diabetic neuroarthropathy was observed in four patients; these are the first cases of this nature reported in the Canadian medical literature. The criteria for this diagnosis included: (1) long-standing diabetes; (2) arthropathy, most frequently involving the foot, which shows deformity, shortening and ulceration without evidence of infection or peripheral circulatory failure; (3) abolition or diminution of pain on weight-bearing; (4) diabetic peripheral neuropathy with impaired sense of position or vibration and weak or absent deep tendon reflexes. Radiographic findings were similar to those in patients with Charcot''s arthropathy from any cause.Tabes dorsalis, leprosy, syringomyelia, myelodysplasia and the arthropathies of corticosteroid therapy were ruled out in these cases. In addition to conventional medical therapy the patients were treated by means of walking-casts for several months.Diabetic neuroarthropathy is probably more common than the medical literature would indicate. Diminished sensation in the lower limbs in diabetics of long standing appears to be the major factor contributing to this disorder.