Catabolism of myo-Inositol to Precursors Utilized for De Novo Glycerolipid Biosynthesis

Systemic injection of [2-3H]myo-inositol into frogs resulted in the incorporation of more than half of the label into glycerolipid classes other than phosphoinositides in retinal rod outer segment membranes. Following methanolysis and differential extraction of isolated lipid classes, radioactivity was recovered primarily in the aqueous phase. After phospholipase C hydrolysis of the total membrane lipids, 97% of the radioactivity was extractable with organic solvents, and 70% of the label in lipids was in 1,2-diglycerides. These results indicate that the label was incorporated primarily into the glyceryl moiety of the membrane glycerolipids. Intraocular injection of frog eyes or in vitro incubation of frog retinas with [2-3H]myo-inositol resulted in the incorporation of radioactivity almost exclusively into phosphoinositides in rod outer segment membranes. Incubation of retinas with [U-14C]glucuronic acid did not result in the formation of labeled retinal lipids. These results suggest that myo-inositol can be catabolized systemically to precursors utilized for glycerolipid biosynthesis in the retina.     

Cloning and Characterization of Yeast LEU4, One of Two Genes Responsible for α-Isopropylmalate Synthesis

By complementation of an alpha-isopropylmalate synthase-negative mutant of Saccharomyces cerevisiae (leu4 leu5), a plasmid was isolated that carried a structural gene for alpha-isopropylmalate synthase. Restriction mapping and subcloning showed that sequences sufficient for complementation of the leu4 leu5 strain were located within a 2.2-kilobase SalI-PvuII segment. Southern transfer hybridization indicated that the cloned DNA was derived intact from the yeast genome. The cloned gene was identified as LEU4 by integrative transformation that caused gene disruption at the LEU4 locus. When this transformation was performed with a LEU4fbr LEU5 strain, the resulting transformants had lost the 5',5',5'-trifluoro-D,L-leucine resistance of the recipient strain but were still Leu+. When it was performed with a LEU4 leu5 recipient, the resulting transformants were Leu-. The alpha-isopropylmalate synthase of a transformant that carried the LEU4 gene on a multicopy plasmid (in a leu5 background) was characterized biochemically. The transformant contained about 20 times as much alpha-isopropylmalate synthase as wild type. The enzyme was sensitive to inhibition by leucine and coenzyme A, was inactivated by antibody generated against alpha-isopropylmalate synthase purified from wild type and was largely confined to the mitochondria. The subunit molecular weight was 65,000-67,000. Limited proteolysis generated two fragments with molecular weights of about 45,000 and 23,000. Northern transfer hybridization showed that the transformant produced large amounts of LEU4-specific RNA with a length of about 2.1 kilonucleotides. The properties of the plasmid-encoded enzyme resemble those of a previously characterized alpha-isopropylmalate synthase that is predominant in wild-type cells. The existence in yeast of a second alpha-isopropylmalate synthase activity that depends on the presence of an intact LEU5 gene is discussed.     

Visual fields of orb web and single line web spiders of the family Uloboridae (Arachnida,Araneida)

Summary In the family Uloboridae, web reduction is associated with changes in web monitoring posture and prosomal features. A spider must extend its first pair of legs directly forward to monitor the signal line of a reduced web. This posture is facilitated by shifts in prosomal musculature that cause reduced web uloborids to have a narrower anterior prosoma, a reduced or absent anterior eye row, and prominent posterior lateral eye tubercles. The eye tubercles and larger posterior eyes of these uloborids suggest that web reduction may also be accompanied by ocular changes that compensate for reduction of the anterior eyes by expanding the visual fields of the posterior eyes. A comparison of the visual fields of the eight-eyed, orb web species Octonoba octonaria and a four-eyed, reduced web Miagrammopes species was made to determine if this is true. Physical and optical measurements determined the visual angles of each species' eyes and the pattern of each species' visual surveillance. Despite loss of the anterior four eyes, the Miagrammopes species has a visual coverage similar to that of O. octonaria. This is due to (1) an increase in the visual field of each of the four remaining Miagrammopes eyes, accruing from an extension of the retina and an increase in the lens' rear radius of curvature, and (2) a ventral shift of each visual axis, associated with the development of an eye tubercle and an asymmetrical expansion of the retina. Miagrammopes monitor their simple webs from twigs or moss where they are vulnerable to predation. Therefore, maintenance of visual cover may enable them to detect predators in time to assume or maintain their characteristic, cryptic posture. It may also allow them to observe approaching prey and permit them to adjust web tension or prepare to jerk their webs when prey strikes.     

Interaction of nitrogenase with nucleotide analogs of ATP and ADP and the effect of metal ions on ADP inhibition

The interaction of a large number of ATP and ADP analogs with nitrogenase from Azotobacter vinelandii, Klebsiella pneumoniae, and Clostridium pasteurianum has been examined. Only 1,N6-etheno-ATP and 2'-deoxy-ATP served as substrates for acetylene reduction. Other triphosphates including GTP, ITP, 8-Br-ATP, alpha,beta-methylene ATP, beta,gamma-methylene ATP, 6-chloropurine riboside triphosphate, and AMP-PNP were inert, showing less than 50% inhibition at levels up to two- to fivefold greater than ATP. Xanthosine triphosphate behaved simply as a chelator of magnesium, activating the enzyme at low levels but strongly inhibiting at high levels. When nucleotide diphosphates were tested as inhibitors with enzyme from A. vinelandii, GDP, dGDP, and 6-chloropurine riboside diphosphate were ineffective, XDP was three- to fivefold less effective, and dADP and 1,N6-etheno-ADP were about equally as effective as ADP. With enzyme from C. pasteurianum, dADP was twofold less effective than ADP, XDP was fivefold less effective, and IDP and 1,N6-etheno-ADP appeared to be ineffective. Results with enzyme from K. pneumoniae were very similar to those obtained with A. vinelandii. Different metal ions were tested in the presence of both ATP and ADP to determine whether preferential binding to one nucleotide or the other might alter the ADP/ATP ratio needed for 50% inhibition of activity. Magnesium and manganese gave the same ratio, while with Fe and Co, slightly less ADP was required for equivalent inhibition. Nickel appeared to reduce the sensitivity of A. vinelandii nitrogenase to ADP inhibition while increasing that of C. pasteurianum, but both effects were less than twofold. Calcium, strontium, and aluminum ions were inert with enzymes from these organisms. Cd and Zn were also ineffective with K. pneumoniae. Two isomers of ATP beta S were prepared by enzymatic synthesis from ADP beta S. The A form was a more potent inhibitor of A. vinelandii nitrogenase.     

Purification and separation of tomato isolectins by chromatofocusing

Tomato lectin can be rapidly separated from a crude extract of tomato fruit proteins by chromatofocusing. The lectin is recovered from a PBE94 column in two peaks, each with a specific activity comparable to that of lectin purified by affinity chromatography on ovomucoid-Sepharose. Both isolectins consist of a single polypeptide chain (Mr 68,000) and have similar properties.     

Evidence for an oscillator other than luteinizing hormone controlling the secretion of progesterone in cattle

The objective of this study was to quantify and compare the frequencies of pulses in ovarian and systemic concentrations of progesterone, systemic concentrations of luteinizing hormone (LH) and rate of ovarian blood flow. Blood was collected simultaneously from previously implanted catheters in the ovarian venous circulation and jugular vein on Day 12 or 13 of estrous cycles from 4 nonlactating dairy cows. Blood was collected at a rate of 2.5 ml/min for 5 min out of every 10 min over an 8-h period. The mean rate of blood flow in the ovarian artery during the 5-min collection period was estimated by an electromagnetic blood flow transducer. Pulses were observed over time in both ovarian and systemic concentrations of progesterone at frequencies that ranged between 0.625 and 0.875 cycles/h (1.1 to 1.5 h/cycle) among the animals. Only one or two episodes of release of LH were observed during the 8-h period, and transient increases in blood flow to the ovaries were associated temporally with each episode of LH release. The estimated frequencies for release of LH and increased blood flow were the same for each animal and ranged between 0.250 and 0.375 cycles/h. A second cycle with a frequency similar to that for LH was evident in the spectral density functions for ovarian and systemic concentrations of progesterone. This cycle was eliminated when the cycle for LH was removed from the data for progesterone, but the magnitude and frequency of the pulses in progesterone were not affected.(ABSTRACT TRUNCATED AT 250 WORDS)     

INTRAVENOUS NUTRITION—A Clinical Evaluation of a 50 Per Cent Dextrose in Water Solution,Containing 1 mg. of Hydrocortisone per 100 ml

A 50 per cent dextrose in water solution, containing 1 mg. of hydrocortisone per 100 ml., was used successfully in 70 patients for intravenous nutritional maintenance and repletion. There were no adverse systemic effects during or following 216 infusions. The only undesirable local reaction was the rare occurrence of pain in the arm when the concentrated solutions were given too rapidly. Glycosuria was minimal if the infusion rate did not exceed 0.85 gm. of glucose per kilogram of body weight per hour, particularly if 50 units of insulin were added to each 550 ml. bottle of 50 per cent dextrose. In patients without significantly elevated serum potassium content, 30 mEq. of potassium chloride, acetate or phosphate was added to each bottle to prevent hypokalemia.Preliminary observations suggest that this new solution may be given safely intravenously, without need for cutdowns or plastic catheters, if the needle is carefully inserted and well immobilized in the arm vein and the duration of the infusion is not too prolonged. Further studies on the effect of such high caloric supplementation plus protein hydrolysates in parenteral nutritional repletion and maintenance are indicated.