Conformational changes of spin-labeled native and modified phosphorylase B

The phosphorylase B labelled with 2,2,6,6-tetramethyl-piperidine-1-oxyl-4-iodacetamide (phosphorylase I) and with 2,2,6,6-tetramethyl-piperidine-1-oxyl-4-ethylmaleinimide (phosphorylase II) was studied. It was shown that label I is characterized by a greater mobility with respect to the protein as compared to label II. In spin-labelled preparations of phosphorylase B the 1,5--2,0 SH-groups of the enzyme monomer having no effect on the enzyme activity were modified. The effects of AMP, glucose-1-phosphate and glucose-6-phosphate on the EPR spectrum of phosphorylase I were studied. The greatest changes in the spectrum (especially in the high field line) were found to occur in the presence of glucose-6-phosphate. These changes are due to the increase in the degree of anisotropic spin rotation. The experimental and theoretical spectra allowing to determine the correlation time for the protein moiety (tau b = 160 ns) were shown to be similar. The local conformation changes were found to occur in the vicinity of one of the two label-bound SH-groups of phosphorylase I. The EPR spectra demonstrate the S-shaped dependence of mobility of phosphorylase I label on concentration of glucose-6-phosphate (0,1--10 mM). In the presence of AMP no S-shaped dependence is observed. Reduced NaBH4 phosphorylase I does not reveal the S-shaped dependence of the label mobility on concentration of glucose-6-phosphate. The degree of the label immobilization in the apo-phosphorylase I--pyridoxal-5-chloromethylphosphonate complex in the presence of glucose-6-phosphate and AMP is the same as in cholophosphorylase I; however, in contrast to the choloenzyme it does not depend on glucose-6-phosphate (0,1--10,0 mM). The changes in the mobility of the spin label of apophosphorylase I and its complex with the AMP analog--adenosine-5'-chloromethylphosphonate--during the choloenzyme reconstruction by pyridoxalphosphate are indicative of participation of AMP and the phosphate group of AMP in the formation of the enzyme active center.     

Participation of the hippocampus in regulating a pathologic inactive motor reaction in dogs

A local pathological inert motor reaction, involving a forced flexion of the forepaw, was formed after simultaneous stimulation with NaCl solution of a part of the tongue brought out to the cheek and electrical stimulation of the forepaw. The reaction decreased (by 70 to 80%) after electrical stimulation of the dorsal hippocampus (beyond CA3). At the same time the animal's emotional stress diminished, as evidenced by the reduced heart rate and a reduced theta-rhythm in the hippocampus.     

Study of the resistance of Candida guilliermondii to polyene antibiotics

250 Candida guilliermondii strains resistant to the polyene antibiotics nystatin, levorin and amphotericin B were obtained using UV irradiation. When the mutant strains became resistant to one of the polyene antibiotics, their resistance to the other ones changed. Phenotypic analysis showed that the resistance of the strains to polyene antibiotics did not make them susceptible to a rise in osmotic pressure and to a change of the temperature of incubation. Some of the polyene-resistant strains were stained in a medium with methylene blue. Analysis of the sterol composition in the mutants by UV spectroscopy showed that the resistance to polyene antibiotics sometimes involved changes in the sterol composition. Two new UV spectrum types were recorded for the sterols of the mutant strains; they differed from the UV spectrum for the sterols of the parent sensitive strain.     

Phylogeny of the order Rodentia inferred from structural analysis of short retrotransposon B1

A large-scale study of short retroposon (SINE) B1 has been conducted in the genome of rodents from most of the known families of this mammalian order. The B1 nucleotide sequences of rodents from different families exhibited a number of characteristic features including substitutions, deletions, and tandem duplications. Comparing the distribution of these features among the rodent families, the currently discussed phylogenetic relationships were tested. The results of analysis indicated (1) an early divergence of Sciuridae and related families (Aplodontidae and Gliridae) from the other rodents; (2) a possible subsequent divergence of beavers (Castoridae); (3) a monophyletic origin of the group Hystricognathi, which includes several families, such as porcupines (Hystricidae) and guinea pigs (Caviidae); (4) a possible monophyletic origin of the group formed by the remaining families, including six families of mouselike rodents (Myodonta). Various approaches to the use of short retroposons for phylogenetic studies are discussed.     

A new pentanucleotide STR-marker, located in the intron of the ING1 tumor suppressor gene and its allelic polymorphism

DNA samples of unrelated subjects from the Volga-Ural region of Russia were examined to study allele polymorphism of the pentanucleotide repeat (TTGTG)8 localized to an intron of the tumor suppressor gene ING1. STR marker was registered in the EMBL database with the accession number AJ277387. In a sample of 119 individuals, three pentanucleotide alleles consisting of seven, eight, and nine repeated monomers were revealed. The allele frequencies were 0.24, 0.74, and 0.02, respectively. Heterozygosity was 0.45. On the basis of these data, the repeat can be regarded as a polymorphic STR marker for the ING1 gene and used in population and clinical studies.     

Evolutionarily-conserved gene CKAP2,located in region 13q14.3 of the human genome, is frequently rearranged in various tumors

The human CKAP2 gene, which is involved in diffuse large B-cell lymphomas, was localized via screening the GeneBridge 4 somatic cell radiation hybrid panel by means of the polymerase chain reaction (PCR). The CKAP2 gene was mapped between the WI-15460 and WI-3673 markers at the boundary between regions 13q14.3 and 13q21.1, at the distance of 14.39 cR (with 4.8 cR per cM) from the WI-5867 framework marker (lod score > 2.26). The human CKAP2 gene displayed high homology to mouse and rat expressed orthologs, A CKAP2-like sequence was found in human chromosome 14 and assumed to be a pseudogene resulting from duplication and subsequent mutations of the CKAP2 gene on chromosome 13. A possible role of the CKAP2 gene in oncogenesis associated with deletions and rearrangements of region 13q14.3-21.1 is discussed.