B2-like repetitive sequence from the X Chromosome of the American mink (Mustela vison)

In analysis of the repeats from the mink X Chromosome (Chr), we have identified a B2-like repetitive sequence of 195 base pairs (bp) flanked by short direct repeats of 14 bp. It contains regions homologous to the split intragenic RNA polymerase III promoter and a 3 A-rich region followed by an oligo(dA) sequence. A feature of the repeat is the presence of a perfect polypyrimidine tract 22 bp in length absent from the known Alu- and Alu-like sequences. Alignment of the mink B2-like sequence and mouse B2-consensus sequence allowed us to estimate their similarity as 55%. The repeat is present in 1–2×10⁵ copies per mink genome and 2–4×10³ copies per X Chr. In situ hybridization analysis demonstrated a similar distribution pattern of the B2-like repeat along the length of all the mink chromosomes including the X. We also observed the presence of mink B2-like hybridizable sequence in the genomes of other Carnivora species.The nucleotide sequence data reported in this paper have been submitted to EMBL Data Library and have been assigned the accession number X52381 (MVB2RPT).     

The steady-state kinetic mechanism of ATP hydrolysis catalyzed by membrane-bound (Na+ + K+)-ATPase from ox brain IV. Rate constant determination

The expressions for the kinetic constants corresponding to the steady state model for hydrolysis of ATP catalyzed by (Na⁺ + K⁺)-ATPase proposed recently are analyzed with the object of determining the rate constants. The theoretical background for the necessary procedures is described. The results of this analysis are: (1) A small class (four) of rate constants are determined directly by the previously published values of the kinetic constants. (2) For a somewhat larger class of rate constants upper and lower bounds may be established. For several rate constants the upper and lower bounds differ by less than a factor 1.6 (for the ‘(Na⁺ + K⁺)-enzyme’, i.e. the enzyme activity with K⁺ and millimolar substrate concentration) and 1.2 (for the ‘Na⁺-enzyme’, i.e. the activity at micromolar substrate concentrations). (3) Experiments on inhibition by K⁺ of the Na⁺-enzyme at various Mg²⁺ concentrations are reported and analyzed. With the additional assumption that the rate constants governing the addition to ATP of Mg²⁺ is independent of whether or not ATP is bound to an enzyme molecule, a set of consistent values for all the 23 rate constants in the mechanism may be obtained. (4) The values of some rate constants lend further support to the contention discussed in a previous paper that the enzyme hydrolyzes ATP along two kinetically distinct pathways, depending on the presence of K⁺ and on the concentration of substrate, without the necessity of having more than one active substrate site per enzyme unit at any time. (5) The results show that while the two enzyme forms, the ‘Na⁺-enzyme’ E₁ and the “K⁺-enzyme” E₂K, add substrate with (second order) rate constants of the same order of magnitude (differing only by a factor of four in favor of the former), the rate constants for the reverse processes differ by a factor of 100, being largest for the K⁺-enzyme. This is the main reason for the large difference in the Michaelis constants for the two forms reported previously. (6) Compatibility of the model with the well-known rapid dephosphorylation of the phosphorylated enzyme in the presence of K⁺ requires the presence, at non-zero steady state concentration, of an enzyme-potassium-phosphate intermediate, which is acid labile and is therefore not detected as a phosphorylated enzyme using conventional methods.     

Chromosomal locations of two DNA segments that flank ribosomal insertion-like sequences in Drosophila: Flanking sequences are mobile elements

We report the chromosomal locations of two repetitive DNA sequences that flank ribosomal insertion-like sequences in Drosophila melanogaster. The chromocentric region of D. melanogaster contains many copies of sequences that are homologous to type 1 ribosomal insertions. These insertion-like elements are interspersed with other DNA segments that we call flanking sequences. Two distinct flanking sequences derived from the same cloned DNA molecule pDmI 101, the HindIII fragments 101E and 101F, were studied. Whole genome Southern blots with DNA from the D. melanogaster stocks Oregon R (P2), gt-1, and gt-X11 showed complex restriction patterns that differed substantially between the three stocks. This and other data show that flanking sequences are members of diverged repetitive sequence families. In situ hybridization to salivary gland chromosomes of gt-1 and gt-X11 showed that both sequences are homologous to the chromocenter and to about 5 to 8 (101E) or 25 to 30 (101F) euchromatic sites in each stock. Most, if not all, of these sites differed in gt-1 and gt-X11. Both 101E and 101F are homologous to the chromocenter and very few euchromatic bands in D. simulans, but 101F is homologous to numerous bands in D. mauritiana. We conclude that the flanking sequences represented by 101E and 101F are mobile elements within the genome of Drosophila. These two sequences differ in several structural features from mobile DNA elements previously described in this organism.We dedicate this paper to Professor W. Beermann at the occasion of his 60th birthday     

Patterns of ribosomal DNA spacer lengths are inherited

The non-transcribed spacer regions in Xenopus laevis ribosomal DNA vary in length, even within a single nucleolar organizer. The pattern of spacer lengths is sufficiently different from one nucleolar organizer to another to allow the pattern to be used as a genetic marker. We have analyzed the spacer patterns of rDNA² from a total of 50 progeny from three separate matings. Spacer patterns were inherited with no detectable change in all but two cases. The reproducibility of the patterns among siblings and their stable inheritance between generations rule out sudden mechanisms for gene evolution, such as the master-slave model, and support more gradual mechanisms.Two animals out of 50 showed marked changes in their rDNA spacer patterns. It is not possible at present to decide which of several possible mechanisms were responsible for the observed changes.     

Self-association of human erythrocyte phosphofructokinase. Kinetic behaviour in dependence on enzyme concentration and mode of association.

The kinetic behaviour of human erythrocyte phosphofructokinase has been analyzed over a relative wide range of enzyme concentration (0.01 -- 1.7 mug/ml). The kinetic cooperativity which becomes apparent when the enzymic reaction rate is plotted versus the fructose 6-phosphate concentration decreases with increasing enzyme concentration. Simultaneously, a decrease of the half-saturation concentration for fructose 6-phosphate [S]0.5 is observed. Maximum velocity passes through a maximum at increasing enzyme concentrations. Sets of curves representing specific enzymic activity of phosphofructokinase versus enzyme concentration obtained at various fixed concentrations of fructose 6-phosphate and ATP are analyzed. The shapes of these curves are interpreted in terms of an association model of human erythrocyte phosphofructokinase, in which an inactive dimer (Mr 190000) and active multimers of the dimeric form are involved. The conclusion is drawn that the sigmoidal shape of the plots of the enzymic reaction rate versus fructose 6-phosphate concentration is partially caused by a displacement of the equilibrium between different states of association of phosphofructokinase to multimers by this substrate. On the other hand, the inhibition of the enzyme by high concentrations of ATP may be partially caused by a shift of this equilibrium to the state of the inactive dimer.     

Mechanism of the mutagenic action of hydroxylamine. IX. The UV-induced cleavage of the N-O bond in N4-hydroxy-and N4-methoxycytidine and N6-methoxyadenosine.

The principal UV-induced (lambda = 2546nm) reaction of N4-hydroxy- and N4methoxycytidines and N6-methoxyadenosine in neutral aqueous solutions is cleavage of the exocyclic N-O bond with the respective formation of cytidine and adenosine. Quantum yields are 2.8x10(-3) and 2.2x10(-3) M/E for the first two compounds and 9.1x10(-3) M/E for N6-methoxyadenosine.