Cryopreservation of Plasmodium chabaudi I: Protection by glycerol and dimethyl sulfoxide during cooling and by glucose following thawing

Two additives, glycerol and dimethyl sulfoxide (Me₂SO), were investigated for toxic and protective effects for the intraerythrocytic stages of Plasmodium chabaudi. After incubation for 15 min, at 0 °C in Me₂SO and at 37 °C in glycerol, with various concentrations of these additives, half the blood from each treatment was cryopreserved in glass capillary tubes cooled at approximately 3600 °C min^?1 by plunging into liquid nitrogen. Warming was rapid, approximately 12000 °C min^?1, produced by agitation in a water bath at 40 °C for 1 min. The effect of dilution in phosphate-buffered saline (PBS) supplemented with various concentrations (5 to 25% $\text{v}\text{v}$) of glucose was also investigated in conjunction with the two cryoprotectants. Survival of both the frozen and the unfrozen control parasites was assayed by the mean time taken for the parasitemia in groups of five mice to reach a level of 2% following intraperitoneal injection of 10⁶ parasitized erythrocytes into each mouse. Glycerol was toxic at concentrations above 10% $\text{v}\text{v}$ and Me₂SO above approximately 15%. The use of glucose in the recovery medium resulted in a substantial improvement in the survival of frozen and unfrozen parasites previously incubated in either cryoprotectant. The amount of glucose required varied with the concentration of additive used, and optimum survival of cryopreserved parasites was obtaind with 10% $\text{v}\text{v}$ glycerol or 15% $\text{v}\text{v}$ Me₂SO and with 15% $\text{w}\text{v}$ glucose in the diluent medium.     

Trichinella spiralis: genetic basis for differential expression of phase-specific intestinal immunity in inbred mice

Responsiveness of mouse strains after phase-specific immunization with Trichinella spiralis is compared. Two strains ($\text{NFR}\text{N, NFS/N}$) showed strong overall responsiveness. The response type could be characterized in phase-specific terms as: strongly anti-adult, weakly to moderately anti-preadult, and strongly antifecundity. By comparison, congenic mice of the C₅₇B1 $\text{10}\text{Sn}$ background (B10·A, B10·D₂, B10·S, B10·Q) displayed poor total responses that could be characterized as: weakly anti-adult, very weakly anti-preadult, weakly anti-fecundity after preadult immunization, and mixed (weak and strong) after adult immunization. The $\text{C}{}_3\text{H}\text{HeJ}$ mouse appeared to be intermediate between the B10·BR and the $\text{NFR}\text{N}$ strains in overall responsiveness. Genetic determinants of anti-preadult or anti-adult responses of $\text{NFR}\text{N}$ strain mice were dominant over their B10 congenic counterparts as shown in F₁, crosses of $\text{NFR}\text{N}$ × B1O·BR mice. Since the $\text{NFR}\text{N}$ (predominantly H-2^q) and the $\text{NFS}\text{N}$ (H-2^S) are both strong responders, while the B10·Q(H-2^q) and B10·S (H-2^S) are weak, it is suggested that the major genes controlling anti-preadult and anti-adult responses are not linked to the major histocompatibility complex. However, variations in anti-adult immunity and anti-fecundity in the B10 congenic mice (B10·Q and B10·S are the strongest responders) suggest that minor genes linked to the MHC exert some control over these responses. Some evidence was obtained for gene complementation as the F₁ cross of $\text{NFR}\text{N}$ and $\text{NFS}\text{N}$ mice responded more vigorously than the parental lines. We conclude that multiple genes determine anti-T. spiralis intestinal responses in mice. The major genes are unlinked to the major histocompatibility complex whereas several minor genes are linked.     

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.     

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.     

Effects of hydrocortisone on the hypercalcemia and plasma levels of 13,14-dihydro-15-keto-prostaglandin E2 in mice bearing the HSDM1 fibrosarcoma

In mice bearing the prostaglandin-producing HSDM₁ fibrosarcoma, the plasma concentration of 13,14-dihydro-15-keto-PGE₂ was elevated before the development of hypercalcemia, and the magnitude of the rise was greater than that of PGE₂. When hydrocortisone, which inhibits synthesis of PGE₂ by HSDM₁ cells in culture, was administered to tumor-bearing mice, the steroid hormone prevented the rises in plasma PGE₂ metabolite and calcium concentrations. At the dose levels used, hydrocortisone did not inhibit the calcium-mobilizing action of parathyroid hormone $\text{in vivo}$ or the bone resorption-stimulating activity of $\text{PGE}{}_2\text{in vitro}$. These findings are consistent with our hypothesis that the hypercalcemic syndrome in HSDM₁ tumor-bearing mice is due to the secretion of PGE₂ by the tumor.     

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 on drug metabolism of carbaryl and 1-naphthol in the mouse

The effects of carbyl and 1-naphthol on hepatic microsomal drug-metabolizing enzyme systems were investigated. The agents were fed at a level of 25 mmol/kg of feed to groups of young male Swiss-Webster mice for 14-day periods. Body weight was depressed by carbaryl, but not by 1-naphthol. The rates of $\text{in vitro}$ metabolism of aniline and benzphetamine were greater than control rates in livers of mice fed carbaryl, but the rate of $\text{in vivo}$ hydrolysis of the carbamate insecticide Zectran was decreased by carbaryl feeding. Administration of 1-naphthol did not change the rates of $\text{in vitro}$ metabolism of either aniline or benzphetamine. Hepatic microsomal concentrations of cytochromes P-450 and b₅ were increased by carbaryl, but feeding of 1-naphthol did not affect levels of either cytochrome. Radiolabeled pentobarbital disappeared from the blood of carbaryl-fed mice more rapidly than from the blood of control animals, and carbaryl-fed mice slept a shorter period of time than controls following pentobarbital administration. The LD50 of an acute oral dose of carbaryl was increased two-fold by feeding carbaryl for 14 days. It was concluded that carbaryl is a weak inducer of hepatic microsomal drug-metabolizing activity, and that the effects observed are not likely due to 1-naphthol.     

Trichinella spiralis: Characterization and strain distribution of rapid expulsion in inbred mice

Appropriately immunized mice display a response that is biologically equivalent to rat rapid expulsion. Only two inbred strains ($\text{NFR}\text{N}\text{and}\text{NFS}\text{N}$ derived from NIH Swiss mice) have been shown to respond in this manner. Mice of the $\text{Balb}\text{c}$, CBA, $\text{A}\text{He}$, C₃H, SJL, or C₅₇Bl strains are “nonresponders” which require approximately twice as much intestinal exposure (in days) to Trichinella spiralis to elicit a response half as effective. Genetically, the responder is dominant, autosomal, and does not appear to be linked to the MHC. The characteristics of mouse and rat rapid expulsion of T. spiralis are not identical but share these features: initial rejection within 24 hr of challenge; a rejection efficiency >90%, from 1 to 5 weeks after the primary; induction of response does not require exposure to the complete infection; rapid expulsion is immunologically specific for preadults; adult worms are resistant. While a genetic basis for responsiveness exists in mice there is, as yet, no evidence for genetic control in rats. In both mice and rats, rapid expulsion is distinguished from the intestinal hyperreactivity associated with rejection of the primary infection by the kinetics and amplitude of the rejection of transplanted adult worms.     

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.     

Gibberellin A4 produced by Sphaceloma manihoticola, the cause of the superelongation disease of cassava (Manihot esculenta)

Gibberellin A₄ was identified by combined gas chromatography-mass spectrometry in the culture medium of $\text{Sphaceloma}\text{manihoticola}$, a fungus known to cause the “superelongation disease” of cassava $\text{(}\text{Manihot}\text{esculenta}\text{)}$. Gibberellin A₄ was synthesized in ageing cultures and reached concentrations as high as 400 μg/l nutrient broth. After $\text{Gibberella}\text{fujikuroi}\text{,}\text{Sphaceloma}\text{manihoticola}$ is the second phytopathogenic fungus known with certainty to produce an active gibberellin in considerable amounts.     

Interaction of human fibrinogen with divalent metal chlorides

Precipitation of human fibrinogen in 0.15 m NaCl occurred at pH 7.4 (Tris-HCl buffer) when ZnCl₂, CuCl₂, NiCl₂, or CoCl₂ were added beyond their respective critical concentrations. The critical concentrations were about $\text{4 × 10}{}^{\mathrm{?5}}\text{m}\text{ZnCl}{}_2\text{, 6 × 10}{}^{\mathrm{?5}}\text{m}\text{CuCl}{}_2\text{, 3 × 10}{}^{\mathrm{?4}}\text{m}\text{NiCl}{}_2\text{and}\text{1 × 10}{}^{\mathrm{?3}}\text{m}\text{CoCl}{}_2$. At pH 5.8 2-(N-morpholino)-ethane sulphonic acid buffer, the critical concentrations were found only for CuCl₂ and ZnCl₂, and were about $\text{3 × 10}{}^{\mathrm{?5}}\text{and}\text{3 × 10}{}^{\mathrm{?4}}\text{m}$, respectively. CaCl₂ and MgCl₂ were not effective up to $\text{1 × 10}{}^{\mathrm{?2}}\text{and}\text{2 × 10}{}^{\mathrm{?2}}\text{m}$ at pH 7.4 and 5.8, respectively. At pH 7.4, precipitation was better in 0.015 m NaCl than in 0.15 m NaCl for both CuCl₂ and ZnCl₂. Little or no conformational change was indicated on binding Cu²⁺ ions. The fluorescence of tryptophan was quenched only by CuCl₂, while other metal ions (ZnCl₂, NiCl₂, CoCl₂ and CaCl₂) were ineffective as quenchers.     

Neutral salt effects on the velocity and activation volume of the lactate dehydrogenase reaction: Evidence for enzyme hydration changes during catalysis

The effects of different neutral salts on the maximal velocity (V) and activation volume ($\text{Δ}\text{V3}$) of the M₄-lactate dehydrogenase reaction were studied to determine the mechanistic basis of the inhibitory effects of these salts. For salting-in salts (which increase protein group solubility), increasing salt concentrations led to reductions in V and increases in $\text{Δ}\text{V3}$, with the order of salt effectiveness following the Hofmeister (lyotropic) series: KSCN > KI > KBr. A 50% reduction in V was associated with an approximately 17 cm³ mol^?1 increase in $\text{Δ}\text{V3}$ for different concentrations of the same salt and for equal concentrations of different salting-in salts. Salting-out salts were also inhibitory, but no uniform correlation between changes in V and $\text{Δ}\text{V3}$ was observed. The strongly salting-out salt KF decreased $\text{Δ}\text{V3}$ at all concentrations. The weaker salting-out salt K₂SO₄ increased $\text{Δ}\text{V3}$ at concentrations below 0.1 m and decreased $\text{Δ}\text{V3}$ at higher concentrations. KCl increased $\text{Δ}\text{V3}$ as the salt concentration was raised to approximately 0.2 m; further increases in KCl concentration were without effect on $\text{Δ}\text{V3}$. The rate and volume effects of these neutral salts, especially the highly regular covariation in V and $\text{Δ}\text{V3}$ found for salting-in salts, seem difficult to explain in terms of salt-induced changes in the geometry of the active site. We propose instead that these salt effects can all be explained in terms of the energy and volume changes which accompany transfers of protein groups (amino acid side chains and peptide backbone linkages) between the hydrophobic interior of the enzyme and the enzyme-water interface during catalytic conformational changes.     

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.     

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.     

Intraperitoneal injection of zymosan in mice induces pain,inflammation and the synthesis of peptidoleukotrienes and prostaglandin E2

Intraperitoneal injection of zymosan in mice induced rapid extravasation and accumulation of plasma proteins in the peritoneal cavity. Neutrophils began to appear in the peritoneal cavity after a lag period of approximately 3 hours. The injected mice exhibited a pain response (writhing) during the first 30 minutes after injection, but writhing ceased before protein or cell accumulation had reached maximum levels. The injection of zymosan induced synthesis of PGE₂ (measured by RIA) which reached maximum levels of 30 minutes, then declined slowly. Peptido-leukotriene levels (detected by bioassay, RIA and HPLC) increased rapidly after injection, reached a peak within an hour of injection and declined to undetectable levels within 4 hours. The early peptido-LT was predominantly LTC₄, while later, LTE₄ was the major component. LTD₄ levels remained low throughout and no LTB₄ was detected at any time. Indomethacin treatment elevated levels of peptido-LTs, recued PGE₂ levels and inhibited writhing. Phenidone reduced peptido-LT levels. $\text{In}$$\text{vitro}$ studies demonstrated that zymosan stimulates LTC4 synthesis by peritoneal cells whereas LTE₄, LTD₄, LTB₄ or monoHETES were not detectable (using HPLC methods). The source of enzymes responsible for the $\text{in}$$\text{vivo}$ metabolism of LTC₄ to LTD₄ and LTE₄ could not be identified.     

The effects of some cations and anions on spin labeled cytoplasmic membranes of bacillus subtilis

The effects of CaCl₂, MgCl₂, LaCl₃ and some alkali halides on $\text{Bacillus subtilis}$ cytoplasmic membranes were studied using stearic acid spin labels. The results indicate that two mechanisms are operating when these ions interact with $\text{B. subtilis}$ membranes. At low ionic concentrations (0 to 0.1 M) there is direct cation binding to the anionic membranes which in the case of Ca²⁺, Mg²⁺ and La³⁺, confers rigidity on the membranes and reaches saturation when the number of cations present equals the number of anionic sites on the lipids. At high concentrations there is a further effect on the membranes that parallels the known organizing/disorganizing effects that the ions studied have on water structure.     

Maternal estrogen and progesterone levels after hypophysectomy in early pregnancy and in term fetuses or newborn monkeys

Pregnant rhesus monkeys ($\text{Macaca}$$\text{mulatta}$) were hypophysectomized at 8–10 weeks gestation to determine effects on plasma levels of estrone (E₁), estradiol-17β (E₂) and progesterone (P). The first group of monkeys was subsequently fetectomized At 107–114 days. After hypophysectomy there was an initial decrease in maternal peripheral plasma E₂ followed by a rise to preoperative levels within 4–5 weeks. Plasma levels of e₁ and P were not markedly altered. After fetectomy, peripheral estrogen concentrations, especially E₂, declined markedly. In the second experimental series, we have examined the effects of maternal hypophysectomy on levels of E₁, E₂ and P either (1) in both mother and newborn baby or (2) in mother, term fetus and umbilical vein. Groups of hypophysectomized and intact pregnant monkeys (3 each) were delivered by cesarean section at the expected time of parturition. Other hypophysectomized and intact monkeys (2 each) delivered normally. E₂ levels were elevated significantly in plasma of hypophysectomized monkeys at the time of cesarean delivery and in newborn babies of hypophysectomized mothers shortly after parturition. Except for these differences, the maternal hypophysis apparently is not a major factor in the control of E₁, E₂ and P concentrations in pregnant rhesus monkeys.     

Steady-state kinetics of ADP-arsenate and ATP-synthesis in Rhodospirillum rubrum chromatophores

(1) Rates of ATP synthesis and ADP-arsenate synthesis catalyzed by Rhodospirillum rubrum chromatophores were determined with the firefly luciferase method and by a coupled enzyme assay involving hexokinase and glucose-6-phosphate dehydrogenase. (2) V_m for ADP-arsenate synthesis was about 2-times lower than V_m for ATP-synthesis. With saturating [ADP], K(As_i) was about 20% higher than K(P_i). With saturating [anion], K(ADP) was during arsenylation about 20% lower than during phosphorylation. (3) Plots of $\text{1}\text{v}$ vs. $\text{1}\text{[}\text{substrate}\text{]}$ were non-linear at low concentrations of the fixed substrate. The non-linearity was such as to suggest a positive cooperativity between sites binding the variable substrate, resulting in an increased $\text{V}{}_{\mathrm{<mtext>m</mtext>}}\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ ratio. High concentrations of the fixed substrate cause a similar increase in $\text{V}{}_{\mathrm{<mtext>m</mtext>}}\text{K}{}_{\mathrm{<mtext>m</mtext>}}$, but abolish the cooperativity of the sites binding the variable substrate. (4) Low concentrations of inorganic arsenate (As_i) stimulate ATP synthesis supported by low concentrations of P_i and ADP about 2-fold. (5) At high ADP concentrations, the apparent K_i of As_i for inhibition of ATP-synthesis was 2–3-times higher than the apparent K_m of As_i for arsenylation; the apparent K_i of P_i for inhibition of ADP-arsenate synthesis was about 40% lower than the apparent K_m of P_i for ATP synthesis. (6) The results are discussed in terms of a model in which P_i and As_i compete for binding to a catalytic as well as an allosteric site. The interaction between these sites is modulated by the ADP concentration. At high ADP concentrations, interaction between these sites occurs only when they are occupied with different species of anion.     

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.