Crystal structure of the carbon monoxide complex of human cytoglobin

Cytoglobin (Cgb) is a vertebrate heme‐containing globin‐protein expressed in a broad range of mammalian tissues. Unlike myoglobin, Cgb displays a hexa‐coordinated (bis‐hystidyl) heme iron atom, having the heme distal His81(E7) residue as the endogenous sixth ligand. In the present study, we crystallized human Cgb in the presence of a reductant Na₂S₂O₄ under a carbon monoxide (CO) atmosphere, and determined the crystal structure at 2.6 Å resolution. The CO ligand occupies the sixth axial position of the heme ferrous iron. Eventually, the imidazole group of His81(E7) is expelled from the sixth position and swings out of the distal heme pocket. The flipping motion of the His81 imidazole group accompanies structural readjustments of some residues (Gln62, Phe63, Gln72, and Ser75) in both the CD‐corner and D‐helix regions of Cgb. On the other hand, no significant structural changes were observed in other Cgb regions, for example, on the proximal side. These structural alterations that occurred as a result of exogenous ligand (CO) binding are clearly different from those observed in other vertebrate hexa‐coordinated globins (mouse neuroglobin, Drosophila melanogaster hemoglobin) and penta‐coordinated sperm whale myoglobin. The present study provides the structural basis for further discussion of the unique ligand‐binding properties of Cgb. Proteins 2011. © 2011 Wiley‐Liss, Inc.     

Geographical variation of photoperiodic wing form determination and genetic background of reproductive diapause in arrhenotokous populations of <Emphasis Type="Italic">Thrips nigropilosus</Emphasis> Uzel (Thysanoptera: Thripidae) in Japan

To determine the level of inter-population variation in wing form and the induction of reproductive diapause as affected by photoperiod, females of five populations of arrhenotokous Thrips nigropilosus Uzel were reared under four different photoperiodic conditions at 18°C. The experimental populations originated from Nakijin-son, Motobu-cho, and Uruma-shi in Okinawa Island, Amami-shi in Amami-Oshima Island, and Wakayama-shi on Honshu Island. In all five populations, only macropterae were observed when the photoperiod was 13 h light/11 h dark. Brachypterae were observed as the light period was reduced to 11.5 or 12 h in the Wakayama population, and to 10 or 11.5 h in the Nakijin, Motobu, and Uruma populations. Brachypterous females were observed in the Amami population only when the light period was 10 h. With the same photoperiod, only brachypterae were observed in the Wakayama and Uruma populations. Short-day conditions also induced reproductive diapause in the Wakayama population but not in the Amami and Okinawa Island populations. Under 10 h light/14 h dark, almost all hybrids obtained from crossing Wakayama with Amami populations entered reproductive diapause. Some populations in Okinawa Island may have been originated in northern localities.     

Group IVA phospholipase A2 regulates testosterone biosynthesis by murine Leydig cells and is required for timely sexual maturation

In the present paper, we report that PLA2G4A (Group IVA phospholipase A2) is important in the development and function of rodent testes. Interstitial cells of rat testes had high PLA2 (phospholipase A2) activity that was very sensitive to the PLA2G4A-preferential inhibitor AACOCF3 (arachidonyl trifluoromethyl ketone). PLA2G4A protein was expressed primarily in the interstitial cells of wild-type mouse testes throughout maturation. Although Pla2g4a knockout (Pla2g4a-/-) male mice are fertile, their sexual maturation was delayed, as indicated by cauda epididymal sperm count and seminal vesicle development. Delayed function of Pla2g4a-/- mice testes was associated with histological abnormalities including disorganized architecture, swollen appearance and fewer interstitial cells. Basal secretion of testosterone was attenuated significantly and steroidogenic response to hCG (human chorionic gonadotropin) treatment was reduced in Pla2g4a-/- mice compared with their Pla2g4a+/+ littermates during the sexual maturation period. Chemical inhibition of PLA2G4A activity by AACOCF3 or pyrrophenone significantly reduced hCG-stimulated testosterone production in cultured rat interstitial cells. AACOCF3 inhibited forskolin- and cAMP analogue-stimulated testosterone production. These results provide the first evidence that PLA2G4A plays a role in male testes physiology and development. These results may have implications for the potential clinical use of PLA2G4A inhibitors.     

Induction of ovarian maturation and development of eggs,larvae and juveniles of the tonguefish,Cynoglossus abbreviates,reared in the laboratory

Cynoglossus abbreviatus spawns from mid-March to mid-April in the Sea of Shimabara in Kyushu. During the spawning season ovarian maturation was successfully induced by injection of the pituitary homogenate ofHypophthalmichthys molitrix. The dose of the aceton-dried pituitary homogenate was 6.5 mg/kg body weight ofC. abbreviatus. It took about 2 days for ovulation after injection at a water temperature of 14 to 16°C. Artificial fertilizations were accomplished on March 29, 1974 and again on April 7, 1984, using the females matured by hormone injection in the latter case only. The larvae were reared on the rotifers,Artemia nauplii,Tigriopus japonicus and copepods collected from the sea over a period of 113 days in 1974 and 58 days in 1984. The eggs were pelagic, spherical, 1.19–1.23 mm in diameter and had 30–50 oilglobules of 0.068–0.095 mm in diameter, and the perivitelline space was narrow. The incubation period was 90–98 hours at a water temperature of 14 to 16°C. The newly hatched larvae were 3.18–3.45 mm TL and had 61–64 myomeres. The larvae had many melanophores and xanthophores on the body, forming three bands on the caudal region, but were lacking chromatophores on the finfolds. The yolk was completely absorbed when the larvae attained a size of 4.7–5.6 mm TL 8 days after hatching. A single elongated dosai fin ray developed on the head in the 8-day old larvae. The ray was reduced in size as long as the other rays 1 or 2 days after metamorphosis. The rudiment of pectoral fins were found on the both sides of the body in the 2-day old larvae, but two of them disappeared after metamorphosis. A pelvic fin first appeared as a ventral bud just anterior to the gut in the larva of 8.39 mm TL. The full count of 4 rays was observed on the larva of 10.83 mm TL. Metamorphosis began 22 days after hatching when the larvae were 11.20 mm TL. The right eye began to shift the left side of the head at night and reached to the final place after 8.5 hours. It took about 36 hours to complete the metamorphosis, including the eye movement and fusion of the hole in the rostral beak. At the last stage of metamorphosis, the dosal, caudal, anal and ventral fins became confluent. The larvae reached the juvenile stage at a size of 13.5–14.0 mm TL, approximately 28 days after hatchling. The growth of larvae reared in 1974 is expressed by the following equations: Y₁ = 3.448 · 1.0507^x (8≦X≦28) Y₂ = 6.3322 · 1.0275^x (28≦X≦75) where Y is the total length (mm) and X is the number of days after hatching. Growth rate changed after metamorphosis.     

Characterization and kinetic analysis of enzyme–substrate recognition by three recombinant lactococcal tripeptidases

Tripeptidases from Lactococcus lactis subsp. lactis (L9PepTR), L. lactis subsp. cremoris (L6PepTR), and L. lactis subsp. hordniae (hTPepTR) were cloned, overexpressed, purified, and characterized. Although these enzymes contained three to seven naturally occurring amino acid differences, both metal-binding and catalytic sites were highly conserved. The k_cat values of hTPepTR were approximately 1.5- to 2-fold higher than those of L9PepTR, while, for L6PepTR, they were approximately 0.8- to 1.4-times the L9PepTR values. The K_m of tripeptidase from subsp. lactis (L9PepTR) was considerably larger when glycine was the amino acid located at both the N- and C-terminus of the peptide substrate. In addition, the K_m values of L9PepTR increased in the following order for YGG, LGG, FGG, SGG, and α-aminoisobutyrylglycylglycine, while the k_cat/K_m decreased in the same order. These results suggest that the dipole moment and steric hindrance of the N-terminal amino acid side chain may be the most important factors controlling substrate specificity.     

Estimating hominid life history: the critical interbirth interval

Unlike any great apes, humans have expanded into a wide variety of habitats during the course of evolution, beginning with the transition by australopithecines from forest to savanna habitation. Novel environments are likely to have imposed hominids a demographic challenge due to such factors as higher predation risk and scarcer food resources. In fact, recent studies have found a paucity of older relative to younger adults in hominid fossil remains, indicating considerably high adult mortality in australopithecines, early Homo, and Neanderthals. It is not clear to date why only human ancestors among all hominoid species could survive in these harsh environments. In this paper, we explore the possibility that hominids had shorter interbirth intervals to enhance fertility than the extant apes. To infer interbirth intervals in fossil hominids, we introduce the notion of the critical interbirth interval, or the threshold length of birth spacing above which a population is expected to go to extinction. We develop a new method to obtain the critical interbirth intervals of hominids based on the observed ratios of older adults to all adults in fossil samples. Our analysis suggests that the critical interbirth intervals of australopithecines, early Homo, and Neanderthals are significantly shorter than the observed interbirth intervals of extant great apes. We also discuss possible factors that may have caused the evolutionary divergence of hominid life history traits from those of great apes.     

Induction of body distension by operations on the nervous system in larvae of the swallowtall, Papilio xuthus

Removal of the ganglion or severance of the nerve cords at the thorax in mature larvae of the swallowtail, Papilio xuthus, induced systemic distension of the body by swallowing excess air. Such a distension, however, was never induced by simultaneous extirpation of the brain, suboesophageal ganglion, or frontal ganglion, or by severance of the recurrent nerve. Removal of an abdominal ganglion induced distension of the posterior part of the body accompanied by shrinkage of the anterior part. The latter phenomenon appears to be induced by a different mechanism from that of systemic distension.     

Ruminal Fermentation and Sugar Concentrations

The maximal amounts of growth of Selenomonas ruminantium were examined in the media containing various amounts of glucose. The yields of cells per unit weight of glucose are linear functions to glucose concentrations in the ranges between zero to 0.005% and 0.005 to 0.7%, Cell yields per glucose are greater in the former range, indicating greater a-mounts of energy are available per glucose at lower concentrations. Growth responses in lactate media containing various amounts of glucose showed that the preincubation with larger amounts of glucose is inhibitory for the following growth and metabolism of lactate. The organism produces predominantly lactate in the glucose medium. However, volatile fatty acid productions increase when the initial concentrations of glucose become low. Isotopic studies showed that the lactate utilization yielding volatile fatty acids is inhibited by the preceding metabolism of high concentrations of glucose. These results were discussed with regard to normal and abnormal fermentations in the rumen.     

Action of sweet-potato beta-amylase on phosphodextrin of potato starch

The specificity of sweet-potato beta-amylase in the vicinity of the phosphate ester groups was studied by determining the structures of the phosphorylated oligosaccharides (alpha-phosphodextrin and beta-limit-alpha-phosphodextrin) formed by its action on potato starch. The beta-limit-alpha-phosphodextrin was separated by chromatography on Dowex-1 (HCOO^?) resin into three fractions that were distinguishable by the d.p. and by the ratio of d-glucose 6-phosphate residues to total organic phosphate. Each fraction contained linear molecules having one phosphate ester group that was not located at the reducing or non-reducing terminals. The smallest phosphodextrin was 6²-phosphorylmaltotriose. It was deduced that beta-amylase hydrolysed (1→4)-α-d linkages from the non-reducing end until one or two d-glucosyl residues remained attached to the phosphorylated residue, depending on whether there was originally an odd or even number of glucosyl residues on the non-reducing side of the phosphorylated residue.     

Effect of pre-treatment with St John's Wort on nephrotoxicity of cisplatin in rats

An herbal health care supplement, St John's Wort (SJW, Hypericum perforatum) has become widely used in the treatment of depression, and is known to interact with therapeutic drugs. Here we report a preventive effect of SJW on cisplatin nephrotoxicity in rats. Rats were given SJW (400 mg/kg/day, p.o.) for 10 consecutive days, and were injected with cisplatin (5 mg/kg, i.v.) on the day after the final SJW treatment. Cisplatin treatment increased the serum creatinine level, which is an index of nephrotoxicity, to 1.51+/-0.22 mg/dl (mean+/-SE) from 0.28+/-0.05 mg/dl (control) on day 5 after the cisplatin injection. This increase fell significantly to 0.86+/-0.13 mg/dl by pre-treatment with SJW. Cisplatin-induced histological abnormality of the kidney was blocked by pre-treatment with SJW. When SJW was administered for 10 days, the amounts of renal metallothionein (MT) and hepatic multidrug resistance protein 2 (Mrp2) were increased to 164.8+/-13.0% and 220.8+/-39.3% (mean+/-SE) of controls, respectively. GSH levels in the kidney and liver were not changed. Total and free cisplatin concentration in serum was not influenced by SJW treatment. In conclusion, the results suggest that pre-treatment with SJW may diminish cisplatin nephrotoxicity.