Binding of cytochrome c to the cytochrome bc1 complex (complex III) and its subunits cytochrome c1 and b1.

Cytochrome $\text{c}$₁, the electron donor for cytochrome $\text{c}$, is a subunit of the mitochondrial cytochrome $\text{bc}$₁ complex (complex III, cytochrome $\text{c}$ reductase). To test if cytochrome $\text{c}$₁ is the cytochrome $\text{c}$-binding subunit of the $\text{bc}$₁ complex, binding of cytochrome $\text{c}$ to the complex and to isolated cytochrome $\text{c}$₁ was compared by a gel-filtration method under non-equilibrium conditions (a $\text{bc}$₁ complex lacking the Rieske ironsulfur protein was used; von Jagow et al. (1977) Biochim. Biophys. Acta $\text{462}$, 549–558). The approximate stoichiometries and binding affinities were found to be very similar. Binding of cytochrome $\text{c}$ to isolated cytochrome $\text{b}$ which is another subunit of the reductase was not detectable by the gel-filtration method. Further, the same lysine residues of cytochrome $\text{c}$ were shielded towards chemical acetylation in the complexes $\text{c}\text{:}\text{c}$₁ and $\text{c}\text{:}\text{bc}$₁. From this we conclude that the same surface area of cytochrome $\text{c}$ is in direct contact with cytochrome $\text{bc}$₁ and with cytochrome $\text{c}$₁ in the respective complexes and that therefore cytochrome $\text{c}$ is most probably the structural ligand for cytochrome $\text{c}$ in mitochondrial cytochrome $\text{c}$ reductase.     

Effects of 19-hydroxy-prostaglandins on oviductal and uterine motility

The effects of 19-hydroxy-prostaglandins (19-OH-PGs) were tested $\text{in}$$\text{vivo}$ on the rabbit oviduct and uterus and on the rhesus monkey ($\text{Macaca}$$\text{mulatta}$) uterus. The 19-OH-PGEs suppressed spontaneous oviductal and uterine activity in the rabbit. The qualitative effect on the rabbit oviduct of 19-OH-PGEs was similar to that of PGE₂. However, the typical response of the rabbit uterus to PGE₂ was an increase in muscle activity. With regard to the rabbit oviduct, 19(R)-OH-PGE₂ was as potent as PGE₂, but 19(S)-OH-PGE₂ was approximately $\text{1}\text{2}$ as potent as PGE₂. Based on the dose of 19-OH-PGEs usually required to cause a minimal suppression and the dose of PGE₂ required to cause a minimal stimulation of rabbit uterine activity, 19(R)-OH-PGE₂ was twice as potent as PGE₂ while 19(S)-OH-PGE₂ was $\text{1}\text{2}$ as potent as PGE₂. Stimulatory effects on the rabbit oviduct and uterus were observed following administration of 19-OH-PGEs and PGF_2α. The potency on the rabbit oviduct of 19(S)-OH-PGF_2α was about $\text{1}\text{5}$ to $\text{1}\text{10}$ that of PGF_2α; the potency of 19(R)-OH-PGF_2α was about $\text{1}\text{10}$ to $\text{1}\text{20}$ that of PGF_2α. Both 19-OH-PGFs were approximately $\text{1}\text{5}$ to $\text{1}\text{10}$ as potent as PGF_2α on the rabbit uterus. At the doses tested 19-OH-PGFs were inactive on the monkey uterus. Thus, these compounds are at least $\text{1}\text{5}$ as active as PGF_2α. In contrast, 19(R)-OH-PGE₂ had approximately the same potency as PGE₂ in stimulating monkey uterine activity; but 19(S)-OH-PGE₂ was approximately $\text{1}\text{3}$ as potent as PGE₂.     

Synthesis of the four isomers of 5-hydroxy-PGI1

We wish to report here the syntheses of (5$\text{S}$, 6$\text{R}$)-5-hydroxy-, (5$\text{R}$, 6$\text{R}$)-5-hydroxy-, (5$\text{R}$, 6$\text{S}$)-5-hydroxy-, and (5$\text{S}$, 6$\text{S}$)-5-hydroxy-PGI₁ and their methyl ester derivatives. Treatment of (5$\text{R}$, 6$\text{S}$)-5-epoxy- and (5$\text{S}$, 6$\text{R}$)-5-epoxy-PFG_1α methyl esters with acid washed silica gel afforded (5$\text{R}$, 6$\text{R}$)-5-hydroxy- and (5$\text{S}$, 6$\text{S}$)-5-hydroxy-PGI₁ methyl esters; correspondingly, silica promoted cyclization of (5$\text{S}$, 6$\text{S}$)-epoxy- and (5$\text{S}$, 6$\text{R}$)-5-epoxy- PGF₁ methyl esters yielded (5$\text{S}$, 6$\text{R}$)-5-hydroxy- and (5$\text{R}$, 6$\text{S}$)-5-hydroxy- PGI₁ methyl esters. Alternatively, the 5-hydroxyl group was introduced into the PGI₁ skeleton $\text{via}$ reaction of the 5-mercuric halides with sodium borohydride in the presence of oxygen. Stereochemical assignments were based on their mode of synthesis and ¹H nmr shift differences.     

Disaggregation of elongation factor 1 by extracts of Artemiasalina

Extracts of 40 hr $\text{Artemia}$$\text{salina}$ nauplii can convert a heavy form of elongation factor 1 (EF-1_H) to a light species (EF-1_L). The data indicate that a protease in the extracts is responsible for this reaction, and these findings may explain the observation that extracts from $\text{Artemia}$$\text{salina}$ nauplii have only EF-1_L whereas before hatching of the $\text{Artemia}$$\text{salina}$ embryos EF-1_H is the predominant species (Slobin and M?ller [1975] Nature 258, 452–454).     

Reflex ovulation in light-induced persistent estrus (LLPE) rats: Role of sensory stimuli and the adrenals

Penile intromissions have been thought to be the primary stimulus for reflex ovulation in light-induced persistent estrus (LLPE) rats, even though other stimuli also trigger reflex ovulation. To clarify the nature of these noncoital stimuli, intact (nonadrenalectomized) LLPE rats were briefly exposed to a variety of environmental stimuli, other than intromissions, and checked for ova 19–22 hr later. Summary of results (number of rats ovulating/number of rats tested): (A₁) home cage ($\text{3}\text{10}$); (B₁) home cage + vaginal taping ($\text{2}\text{9}$); (C₁) home cage + male-soiled bedding ($\text{15}\text{28}$); (D₁) novel cage ($\text{2}\text{11}$); (E₁) novel cage + vaginal taping ($\text{2}\text{11}$); (F₁) novel cage + vaginal taping + male-soiled bedding ($\text{9}\text{19}$); (G₁) novel cage + vaginal taping + male-soiled bedding + male mounts without intromissions ($\text{14}\text{26}$). The percentage of LLPE rats that ovulated in the last-mentioned test condition was related to the degree of proceptivity/receptivity of the LLPE females. Eight of eight proceptive LLPE females ovulated, but only $\text{6}\text{18}$ nonproceptive females ovulated. To account for reflex ovulation in the absence of intromission it has been suggested that adrenal progesterone (P) stimulates release of an ovulatory quota of luteinizing hormone. This study demonstrates no significant differences in percentage of LLPE females ovulating in corresponding groups of adrenalectomized (ADX) and adrenal-intact females. Summary of results: A₂ = $\text{0}\text{6}$, B₂ = $\text{5}\text{15}$, C₂ = $\text{4}\text{16}$, D₂ = $\text{2}\text{14}$, E₂ = $\text{5}\text{13}$, F₂ = $\text{7}\text{19}$, G₂ = $\text{10}\text{21}$. Conclusion: (a) Exposure to a factor in male-soiled bedding induces reflex ovulation in a significant proportion of adrenal-intact LLPE animals while exposure to a novel cage and/or vaginal taping does not, (b) penile intromissions are not the primary stimulus for reflex ovulation in intact proceptive LLPE rats, and (c) adrenal P is not required for reflex ovulation after tests with noncoital stimuli.     

The influence of dna A and dna C mutations on the initiation of plasmid DNA replication

The dependence of the replication of several plasmids on the chromosome-determined initiation products, $\text{dna}$ A and $\text{dna}$ C, has been studied. The initiation of the replication of $\text{Col}$ E₁ DNA requires the chromosomal $\text{dna}$ A product. In contrast two de-repressed transfer factors ($\text{R 1 drd 16}$ and $\text{Hly}$₁₅₂) seem to determine a corresponding plasmid-specific factor. The $\text{dna}$ C-product is necessary for the ordered initation of all plasmids studied. The addition of low concentrations of chloramphenicol leads to a relaxed replication of $\text{Col}$ E₁ DNA at the restrictive temperature in $\text{dna}$ A-mutants, but not in $\text{dna}$ C-mutants.     

Isolation and properties of the cytochrome B-C1 complex from Rhodopseudomonas sphaeroides

A highly purified and active cytochrome $\text{b}$-$\text{c}$₁ complex has been isolated from the chromatophores of the photosynthetic bacteria $\text{Rhodopseudomonas}\text{sphaeroides}$ R-26, through steps of Triton X-100 solubilization, salt fractionation and calcium phosphate column chromatography. The isolated enzyme complex catalyzes fully antimycin A sensitive oxidation of ubiquinol by cytochrome $\text{c}$ with a turnover number of 1500 per minute at 23° based on cytochrome $\text{c}$₁. It contains 8.3 nmoles of cytochromes $\text{b}$ and $\text{c}$₁ per mg protein and shows four polypeptides in the sodium dodecylsulfate polyacrylamide gel electrophoresis.     

The further investigation of physical and photochemical properties of quasimetacyclophane derived from thymine

The thymine derived quasimetacyclophane exist in two conformers $\text{a}$ and $\text{b}$. The absorption spectra of $\text{a}$ and $\text{b}$ were evaluated and the conformational equilibrium in different solvents /H₂O : EtOH/ were examined. The rate constant k_?1 for reaction $\text{b}\text{→}\text{a}$ was established as well as E_a.     

Synthesis of d1-4,5,6-trinor-3,7-inter-m-phenylene-3-oxaprostaglandins including one which inhibits platelet aggregation

The synthesis of $\text{d1}$-4,5,6-trinor-3,7-inter-$\text{m}$-phenylene-3-oxaprostaglandins oxaprostaglandins of the E₁ and F₁α series⁷ from 6-endo-(1-heptenyl)-bicyclo[3:1:0]hexan-3-one (III), is described. Preliminary biological screening data for gerbil colon smooth muscle stimulation, rat blood pressure and substrate specificity toward 15-hydroxyprostaglandin dehydrogenase is presented. Platelet function studies, both $\text{in vitro}$ and $\text{in vivo}$ of $\text{d1}$-4,5,6-trinor-3,7-inter-$\text{m}$-phenylene-3-oxa-PGE₁, methyl ester (VIII) are presented.     

Crystallography of Bacillus subtilis levansucrase

Levansucrase, an exocellular enzyme, has been isolated from a high producer mutant, the BS5C4 constitutive strain, of Bacillus subtilis. Three crystalline forms have been obtained, all three belonging to the orthorhombic space group P2₁2₁2₁. The most suitable form for a three-dimensional structure investigation has cell dimensions, $\text{a = 68}\text{A}\text{?}$, $\text{b = 125}\text{A}\text{?}$, $\text{c = 54}\text{A}\text{?}$. There is one molecule in the asymmetric unit.     

Time resolved spectroscopy of the tryptophyl fluorescence of the E. coli LAC repressor.

A new crystal form of a mitogenic lectin from pea seeds ($\text{Pisum sativum}$) has been obtained which is suitable for high resolution structural work. The crystals are orthorhombic, space group P2₁2₁2₁, with unit cell dimensions: $\text{a}$ = 64.2Å, $\text{b}$ = 72. 7Å, $\text{c}$ = 108. 3Å. The asymmetric unit contains one protein molecule.     

Isolation of QP-C and reconstitution of the QH2-c reductase

A ubiquinone protein, QP-C, which acts in the cytochrome $\text{b}\text{-}\text{c}$₁ region has been solubilized. The isolated QP-C shows one band of molecular weight 15,000 in polyacrylamide gel electrophoresis in sodium dodecyl sulfate and isoelectric focusing at the isoelectric point of pH 3.6. QP-C reconstitutes with the QP-C deficient $\text{b}\text{-}\text{c}$₁ complex to restore the OH₂-cytochrome $\text{c}$ activity and recover the EPR signal of the complex.     

Stimulation by interferon of induction of differentiation of human promyelocytic leukemia cells

F₁-ATPase was isolated from yeast $\text{S.}\text{cerevisiae}$. The constituent subunits 1 and 2 were purified by gel permeation chromatography, and their amino acid compositions determined. Both subunits have a similar composition except for $\text{1}\text{2}$ cystine, methionine, leucine, histidine, and tryptophan. When F₁ is treated for three hours with 5′-p-[³H]fluorosulfonylbenzoyl adenosine in dimethylsulfoxide, 90% of the activity is lost. Disc gel electrophoresis of the modified complex showed that over 90% of the label was associated with subunit 2. A labelled peptide from a $\text{S.}\text{aureus}$ digest of subunit 2 was isolated and sequenced. It had the following amino acid sequence: His-Try¹-Asp-Val-Ala-Ser-Lys-Val-Gln-Glu, whereby Tyr¹ is the modified amino acid residue. This sequence shows homology to other sequences obtained from maize, beef heart, and $\text{E.}\text{coli}$ F₁-ATPases.     

Protein synthesis in rabbit reticulocytes: control of protein synthesis initiation by a met-tRNA Met f deacylase and peptide chain initiation factors

The 0.5M KCl wash of rabbit reticulocyte ribosomes (I fraction) catalyzes the deacylation of Met-tRNA_f^Met. Upon DEAE-cellulose column chromatography, the deacylase activity elutes with the 0.1M KCl wash of the column (f1) and is well-resolved from the peptide chain initiation factors (1–3). The deacylase activity is specific for Met-tRNA_f^Met (retic., $\text{E.}$$\text{coli}$). Other aminoacyl tRNAs tested including fMet-tRNA_f^Met (retic., $\text{E.}$$\text{coli}$), Phe-tRNA ($\text{E.}$$\text{coli}$), Val-tRNA (retic.), and Arg-tRNA (retic.) are completely resistant to the action of the deacylase. In the presence of the peptide chain initiation factor (IF1) and GTP, retic. Met-tRNA_f^Met forms the initiation complex Met-tRNA_f^Met:IF1:GTP (2), and in this ternary complex Met-tRNA_f^Met is not degraded by the deacylase. $\text{E.}$$\text{coli}$ Met-tRNA_f^Met binds to IF1 independent of GTP, and in this complex, this Met-tRNA_f^Met is degraded by the deacylase.Prior incubation of f1 with Met-tRNA_f^Met (retic.) strongly inhibited protein synthesis initiation, presumably due to deacylation of the initiator tRNA. This inhibition by f1 was completely prevented when Met-tRNA_f^Met (retic.) was pre-incubated with peptide chain initiation factors.     

Crystallographic analysis of the phytoagglutinin from Abrus precatorius by X-ray diffraction and electron microscopy

The lectin from the seeds of Abrus precatorius has been crystallized and the crystals subjected to study by X-ray diffraction and electron microscopy. Three closely related crystal forms were obtained, of orthorhombic space group P2₁2₁2₁ with $\text{a = 138}\text{A}\text{?}$, $\text{b = 142}\text{A}\text{?}$, and $\text{c = 173}\text{A}\text{?}$, of tetragonal space group P4₁2₁2 with $\text{a = b = 136}\text{A}\text{?}$, $\text{c = 176}\text{A}\text{?}$, and a twinned intermediate of the first two. From electron microscopy and two-dimensional spatial filtering of electron micrographs of the crystals, the molecule appears to consist of four similar domains grouped in a roughly planar diamond-shaped arrangement having a local intramolecular dyad axis. The average diameter of the Abrus lectin molecule is 50 to 60 Å and the individual domains appear to have a diameter of about 25 Å.     

Structure of the "keratosulfate-like" material in liver from a patient with G M1 -gangliosidosis ( -D-galactosidase deficiency)

Large amounts of a glycopeptide containing galactose, $\text{N}$-acetylglucosamine, $\text{N}$-acetylgalactosamine and threonine in the ratio 4:3:1:1, together with smaller amounts of mannose, fucose, sialic acid, sulfate, serine, and other amino acids were isolated from the liver of a patient with G_M1-gangliosidosis. Treatment with mild alkali and sodium borohydride indicated an $\text{O}$-glycosidic linkage between $\text{N}$-acetylgalactosamine and threonine. All the hexosamine residues were resistant to sodium metaperiodate whereas 2 out of 4 D-galactose residues were destroyed. Further studies indicated that one of the galactose residues was 1→3 linked to $\text{N}$-acetylgalactosamine (as in G_M1) and the other 1→4 linked to $\text{N}$-acetylglucosamine as found in skeletal keratosulfate.     

Fever induced in marine arthropods by prostaglandin E1

Injections of prostaglandin E₁ into the haemocoel of the marine arthropods $\text{Limulus}$$\text{polyphemus}$, $\text{Homarus}$$\text{americanus}$ and $\text{Penaeus}$$\text{duorarum}$ induced a behaviorally mediated fever (increase in preferred temperature, resulting in increased body temperature). The finding that PGE₁ is pyorgenic to arthropods as well as to mammals suggests that arthropods can be used as simple experimental models for further investigations of the neuropharmacological role of prostaglandins in fever.     

Trichinella spiralis: Genetics of worm expulsion in inbred and F1 mice infected with different worm doses

The nematode Trichinella spiralis is rejected from the intestine at a time that is characteristic for each inbred strain of mouse. Previous work (R. G. Bell et al. 1982a) had empirically identified strong, intermediate, and weak phenotypes (NFR, $\text{C}\text{H}\text{He}$, and $\text{C57}\text{10}$ mice, respectively) in mice infected with 400 muscle larvae. It is shown that this classification applies to another eight inbred strains: SWR, $\text{DBA}\text{2}$, $\text{DBA}\text{1}$, LP, $\text{Bub}\text{Bn}$—all intermediate, and $\text{NZB}\text{BIN}$, C57L, A, and Mus molossinus—all weak. This phenotypic classification consistently applies with infections of 400–800 muscle larvae. Below doses of 300 muscle larvae, the strain designation of phenotype does not consistently apply. By this it is meant that the relative rejection rate changes for certain strains so that eventually some strains that were strong (NFR) or intermediate (AKR) responders to 400 muscle larvae become weak responders to 50 muscle larvae. Other strains increase their relative rejection time (B10 · BR, B10 · Q) while many do not change (NFS, $\text{C}\text{3}\text{Heb}\text{Fe}$, $\text{DBA}\text{2}$, $\text{DBA}\text{1}$). The phenomenon is most apparent in inbred parental strains rather than in F₁ crosses, and it represents a phenotypic variation in rejection time that is dependent on dose. It is also demonstrated that time of rejection is directly proportional to dose in all inbred and F₁ mouse strains that we have examined. Analysis of F₁ crosses shows that most have the rejection time of the strongest responding parental line, suggesting simple genetic control of strong, intermediate, and weak responses. Two F₁ crosses invalidated this theory. The $\text{DBA}\text{1}\text{×}\text{C3}\text{H}\text{He}$ (intermediate × intermediate) showed a strong response. The additive effects of parental rejection phenotype indicated that these lines could not be genetically identical for intermediate responsiveness. Similarly, the NFR (strong) × B10 · BR (weak) F₁ showed intermediate rejection, indicating partial dominance of $\text{C}\text{57}\text{B}\text{1}\text{10}$ genes over the strong responder NFR strain. Neither the primary expulsion time phenotype, phenotypic variation to low doses, or the rejection characteristics of F₁ crosses could be ascribed to genes linked to the major histocompatibility complex.     

Neocarzinostatin chromophore: Presence of a cyclic carbonate subunit and its modification in the structure of other biologically active forms

On the basis of spectroscopic evidence, opening of a five-membered cyclic carbonate ring (1,3-dioxolan-2-one) in the C₁₅-subunit of the previously determined partial structure $\text{1}$ (Fig. 1) of the major neocarzinostatin chromophore (NCS-Chrom $\text{A}$), is proposed to account for its base-catalyzed methanolysis to NCS-Chrom $\text{C}$. NCS-Chrom $\text{B}$, apparently an authentic natural product present as a minor component in all preparations of NCS studied, was found to be formally equivalent to the hydrolysis/decarboxylation product of the cyclic carbonate functionality in NCS-Chrom $\text{A}$. The mercaptan-dependent DNA strand-scission activity, equivalent for NCS-Chrom $\text{A}$, $\text{B}$ and $\text{C}$, is independent of the integrity of the cyclic carbonate ring system and implicates a secondary site in the C₁₅-substructure for mercaptan activation.     

Proton and carbon magnetic resonance studies of the synthetic polypentapeptide of elastin.

Proton and ¹³C magnetic resonance studies are reported on the synthetic polypentapeptide of elastin, HCO-(Val₍₁₎-Pro₍₂₎-Gly₍₃₎-Val₍₄₎-Gly₍₅₎)_n-Val-OMe, where n ∼- 18. Temperature and solvent dependence of peptide N$\text{H}$ chemical shift and solvent dependence of peptide carbonyl chemical shift were used to delineate these moieties preliminary to identification of secondary structure.Based on these studies it is proposed, for the organic solvents of dimethyl sulfoxide, methanol, and low-temperature trifluoroethanol, that dynamic hydrogen bonds form in order of decreasing frequency of occurrence between the Val₍₁₎$\text{C}$O and the Val₍₄₎ N$\text{H}$ (a β-turn), between the Gly₍₃₎ N$\text{H}$ and the Gly₍₅₎$\text{C}$O (an 11-atom, hydrogen-bonded ring), and a more limited interaction between the Gly₍₃₎$\text{C}$O and the Gly₍₅₎ N$\text{H}$ (a γ-turn).Arguments are presented that relate the conformational features proposed above to the coacervate, which is a filamentous state.