Characterization of the nucleotide tight-binding sites of the isolated mitochondrial F1-ATPase

The properties of the nucleotides tightly bound with mitochondrial F1-ATPase were examined. One of three bound nucleotide molecules is localized at the site with Kd approximately 10(-7) M and released with koff approximately 0.1 s-1. The second nucleotide molecule is bound with the enzyme with Kd approximately 10(-8) M and koff for its dissociation is 3 X 10(-4) s-1. The third is never released even in the presence of 1 mM ATP or ADP. The last two nucleotides are believed to be bound at the noncatalytic sites of F1-ATPase. Pyrophosphate promotes liberation of two releasable nucleotide molecules, decreasing the affinity of the enzyme to AD(T)P. From the results obtained it follows that the only suitable criterion for localization of the nucleotide at the F1-ATPase catalytic site is the high rate (koff greater than or equal to 0.1 s-1) of its spontaneous release.     

DNA polymorphism of human HLA-linked complement C4 allotypes, including C4 null alleles, in the Finnish population

Human HLA-linked complement C4 gene products, C4A and C4B, show extensive genetic polymorphism. In both loci, an allele without a gene product, C4 null, is also observed. We have performed a restriction enzyme analysis of genomic DNA samples from individuals having all common (frequency over 1%) C4 protein allotypes observed in the Finnish population. Only one allotype-specific RFLP marker was observed. With some enzymes a DNA polymorphism was observed, which was not detectable by C4 protein typing. Analysis of 10 different C4B null haplotypes and 4 C4A null haplotypes suggested that only one haplotype, HLA-B8 C4A0 B1, carried a C4A gene deletion. This was observed in all 4 unrelated individuals homozygous for this haplotype.     

DNA polymerase epsilon: the latest member in the family of mammalian DNA polymerases

DNA polymerase epsilon is a mammalian polymerase that has a tightly associated 3'----5' exonuclease activity. Because of this readily detectable exonuclease activity, the enzyme has been regarded as a form of DNA polymerase delta, an enzyme which, together with DNA polymerase alpha, is in all probability required for the replication of chromosomal DNA. Recently, it was discovered that DNA polymerase epsilon is both catalytically and structurally distinct from DNA polymerase delta. The most striking difference between the two DNA polymerases is that processive DNA synthesis by DNA polymerase delta is dependent on proliferating cell nuclear antigen (PCNA), a replication factor, while DNA polymerase epsilon is inherently processive. DNA polymerase epsilon is required at least for the repair synthesis of UV-damaged DNA. DNA polymerases are highly conserved in eukaryotic cells. Mammalian DNA polymerases alpha, delta and epsilon are counterparts of yeast DNA polymerases I, III and II, respectively. Like DNA polymerases I and III, DNA polymerase II is also essential for the viability of cells, which suggests that DNA polymerase II (and epsilon) may play a role in DNA replication.     

FGFR-4, a novel acidic fibroblast growth factor receptor with a distinct expression pattern.

We have previously identified two novel members of the fibroblast growth factor receptor (FGFR) gene family expressed in K562 erythroleukemia cells. Here we report cDNA cloning and analysis of one of these genes, named FGFR-4. The deduced amino acid sequence of FGFR-4 is 55% identical with both previously characterized FGFRs, flg and bek, and has the structural characteristics of a FGFR family member including three immunoglobulin-like domains in its extracellular part. Antibodies raised against the carboxy terminus of FGFR-4 detected 95 and 110 kd glycoproteins with a protein backbone of 88 kd in COS cells transfected with a FGFR-4 cDNA expression vector. The FGFR-4 protein expressed in COS cells could also be affinity-labeled with radioiodinated acidic FGF. Furthermore, ligand binding experiments demonstrated that FGFR-4 binds acidic FGF with high affinity but does not bind basic FGF. FGFR-4 is expressed as a 3.0 kb mRNA in the adrenal, lung, kidney, liver, pancreas, intestine, striated muscle and spleen tissues of human fetuses. The expression pattern of FGFR-4 is distinct from that of flg and bek and the yet additional member of the same gene family, FGFR-3, which we have also cloned from the K562 leukemia cells. Our results suggest that FGFR-4 along with other fibroblast growth factor receptors performs cell lineage and tissue-specific functions.     

Vitamin E Status of Healthy Swedish Cattle*

Using high pressure liquid chromatography the serum concentration of vitamin E was measured in dairy cows fed either hay or silage as their main roughage, in calves fed milk-replacer, and in young intensively fed bulls. The concentrates fed to the cows, calves and bulls were supplemented with 5–10, 25 and 5–10 mg DL-α-tocopheryl acetate per kg, respectively, and the milk-replacer for the calves was supplemented with 50 mg DL-α-tooopheryl acetate per kg powder. Cows fed silage as their main roughage had higher serum vitamin E concentrations ( $${\rm{\bar x}}$$ : 3.8–5.2 mg/l) than cows fed only hay ( $${\rm{\bar x}}$$ : 2.5–4.1 mg/1). Lactating cows had higher vitamin E concentrations than dry cows ( $${\rm{\bar x}}$$ : 4.1–5.2 and 2.5–3.8 mg/l, respectively) and calves and bulls had much lower vitamin E concentrations ( $${\rm{\bar x}}$$ : 1.4 and 1.2 mg/l, respectively) than cows. Thirty per cent of the calves and 41 % of the bulls had serum vitamin E concentrations less than 1.0 mg/l, suggesting that in these animals the conventional level of supplementation of feeds with DL-α-tocopheryl acetate in Sweden is probably inadequate for the prevention of nutritional muscular degeneration and other negative effects.     

Strain while skiing and hauling a sledge or carrying a backpack

Eight soldiers on skis transported three loads of different weights on the level, uphill and downhill. The load was placed either on a cargo sledge or in a backpack or divided between the sledge and the backpack. The sledge had a new type of haulage-shaft, which was fixed to both sides of the pelvis. A service belt spread the pull over the whole upper body. The physical stain of different transport methods and the serviceability of the sledge was studied by measuring heart rate (HR), oxygen consumption, ventilation, and perceived exertion. The results indicate that both absolute and relative strain were systematically lower when pulling the load on the sledge than when carrying it in the backpack and on the sledge. HR when pulling a load equal to the human body weight on the sledge was on average 133 beats X min-1; HR was significantly higher 144 beats X min-1 when the load was divided between backpack and sledge. At the lower load level the differences between the transport methods were not significant for HR, oxygen consumption or ventilation. Uphill travel increased oxygen consumption by about 50% over that on the level. Perceived exertion at all load levels was significantly lower with the sledge than with the backpack alone or in combination. The estimated maximal allowable working time emphasized the advantage of the sledge and the importance of high physical working capacity. The maneuverability of the sledge with the new haulage shaft was good and the braking mechanisms worked well.(ABSTRACT TRUNCATED AT 250 WORDS)     

Chloroplast DNA variation inAnemone s. lato (Ranunculaceae)

Chloroplast DNA of seven species belonging toAnemone (sectt.Omalocarpus, Anemonidium, andAnemonanthea),Hepatica, andPulsatilla have been analyzed by restriction enzymes. According to the dendrogram constructed, the sections ofAnemone and the generaHepatica andPulsatilla seem to be evolutionary approximately equidistant to each others. This supports the concept that these groups should be treated on a similar taxonomic level, either as genera or subgenera.     

An approach to mapping haplotype-specific recombination sites in human MHC class III

Studies of the major histocompatibility complex (MHC) in mouse indicate that the recombination sites are not randomly distributed and their occurrence is haplotype-dependent. No data concerning haplotype-specific recombination sites in human are available due to the low number of informative families. To investigate haplotype-specific recombination sites in human MHC, we here describe an approach based on identification of recombinant haplotypes derived from one conserved haplotype at the population level. The recombination sites were mapped by comparing polymorphic markers between the recombinant and assumed original haplotypes. We tested this approach on the extended haplotype HLA A3; B47; Bf ^* F; C4A ^* 1; C4B ^* Q0; DR7, which is most suitable for this analysis. First, it carries a number of rare markers, and second, the haplotype, albeit rare in the general population, is frequent in patients with 21-hydroxylase (21OH) defect. We observed recombinants derived from this haplotype in patients with 21OH defect. All these haplotypes had the centromeric part (from Bf to DR) identical to the original haplotype, but they differed in HLA A and B. We therefore assumed that they underwent recombinations in the segment that separates the Bf and HLA B genes. Polymorphic markers indicated that all break points mapped to two segments near the TNF locus. This approach makes possible the mapping of preferential recombination sites in different haplotypes.     

Significance of Ca2+ and K+ in Micrasterias Growth and Morphogenesis

Significance of Ca²⁺ and K⁺ for the complex morphogenesis ofMicrasterias, which takes place through multipolar tip growth,was investigated. Studies with different external Ca²⁺ concentrationsand Ca²⁺ channel inhibitors LaCl₃ and verapamil indicate thatCa²⁺ and Ca²⁺ channels are essential in the development, whiletreatments with different K⁺ concentrations and K⁺ channel inhibitorTEA demonstrate that potassium or K⁺ channels are not neededin the process, albeit the existence of K⁺ channels. K⁺ is notneeded even for the regulation of turgor pressure, which wasfound to decrease clearly during cell development. The plasmamembrane ATPase inhibitors diethylstilbesterol (DES) and Na-orthovanadatestop morphogenesis and indicate the importance of ion pumpsin the developmental process. Both supraoptimal, external K⁺and Ca²⁺ cause abundant Ca²⁺ precipitate formation in chloroplasts,which shows that chloroplasts are important in regulation ofcytoplasmic Ca²⁺ metabolism and that K⁺ activates the uptakeof Ca²⁺ through Ca²⁺ channels. (Received June 13, 1995; Accepted September 13, 1996)