Thermal melting and circular dichroism of DNA complexes with (Lys-Ala-Ala)10 and (Lys-Ala-Ala)n: effect of polypeptide chain length

DNA complexes with polypeptides (Lys-Ala-Ala)₁)] and (Lys-Ala-Ala)₃₄ have been studied using the methods of thermal melting and circular dichroism. Derivative melting curves of (Lys-Ala-Ala)₁₀ DNA differed substantially from those of (Lys-Ala-Ala)₃₄ prepared either by salt gradient dialysis or by direct mixing. Melting curves of the former complex were unimodal or bimodal with T_m increasing continuously withn input lysin-to-DNA phosphate ratio (r); those of the latter complex consisted of three separate transitions with T_m values almost independent of r. Complete reversibility of binding in the (Lys-Ala-Ala)₁₀-DNA system but a slow redistribution of (Lys-Ala-Ala)₃₄ on DNA at low temperature were found in the redistribution experiments Much faster redistribution from denatured to native DNA occurs at the temperature of melting, contributing to the unusual trimodal melting pattern. Circular dichroism curves are very similar for both complexes and indicate little change of DNA conformation upon polypeptide binding.     

Characterization of a novel 14 kDa bile acid-binding protein from rat ileal cytosol

A 14 kDa polypeptide in rat ileal cytosol has been identified as the major intestinal cytosolic bile acid-binding protein (I-BABP) by photoaffinity labeling with the radiolabeled 7,7-azo derivative of taurocholate (7,7-azo-TC). To further characterize I-BABP, the protein was purified by lysylglycocholate Sepharose 4B affinity and DE-52 anion-exchange chromatography. The purified I-BABP contained a single 14 kDa band on SDS-PAGE. The 14 kDa protein showed a 26-fold increase in binding affinity for [3H]7,7-azo-TC compared to cytosolic protein. Immunoblotting of protein fractions separated by affinity chromatography showed that neither liver fatty acid binding protein (L-FABP) nor intestinal fatty acid binding protein (I-FABP) bind to the affinity column and that the 14 kDa protein which bound to the column and was subsequently eluted with detergent did not cross-react with anti-L-FABP or anti-I-FABP. The 14 kDa protein labeled with [3H]7,7-azo-TC was radioimmunoprecipitated from cytosol by rabbit antiserum raised against purified I-BABP. I-BABP was shown to have a blocked N-terminus; however, its mixed internal sequence generated from cyanogen bromide-cleaved protein and amino acid composition indicated that it was related to (although clearly distinct from) both I-FABP and L-FABP. These studies have isolated a 14 kDa bile acid-binding protein from rat ileal cytosol which is immunologically and biochemically distinct from I-FABP and L-FABP.     

Atom ordering in cuboctahedral Ni-Al nanoalloys

Energy calculations have been carried out on high-symmetry cuboctahedral Ni-Al nanoalloy clusters, of varying composition, with the interatomic interactions modelled by the Gupta many-body potential. Relaxations of cuboctahedral fragments cut from the bulk lattice of Ni₃Al, with 13-561 atoms, were undertaken, as were relaxations of high symmetry clusters with 55 and 147 atoms. The lowest energy isomers were found to be dominated by three factors: the tendency toward mixing due to the favourable energy of mixing, Δ_mixE; the size difference between nickel and aluminium; and the higher cohesive and surface energy of nickel compared to aluminium. The latter two factors favour Al-segregation to the surface. The most stable Ni:Al composition approaches 3:1 for larger clusters.     

A small one-band paracentric inversion inv (4) (p15.3p16.3).

A neonate with aniridia was found to have a one band paracentric inversion of the short arm of chromosome 4. This was initially difficult to interpret on high resolution banding. The inversion was present in three generations of the family.     

Some meteorological conditions associated with forest fires in Galicia (Spain)

The present paper offers data on the effect of certain meteorological parameters on the outbreak of forest fires in Galicia (Spain). In a day-to-day analysis, the values of the stability of the air column and the saturation deficit at the lower levels are related to the number of fires occurring in the following 24 h.     

Continuous-flow solid-phase synthesis of potential antigenic peptides of hepatitis B virus

The synthesis of three hepatitis B surface antigens derived from S and pre-S proteins (adw S(140-147), [Tyr148] adw S(139-148), and adw pre-S(120-145)) has been accomplished by the continuous flow Fmoc-polyamide solid phase method. The use of different scavengers and trimethylsilyl bromide (TMSBr) in trifluoroacetic acid as deprotecting procedures is discussed.     

Identification of UDP-linked murein precursors as contaminants in recombinant proteins of low molecular weight.

The A(280)/A(260) ratio of a purified protein is frequently used as an indication of the purity of the preparation with respect to nucleic acids. We show here that for low-molecular-weight recombinant proteins purified from Escherichia coli, a low A(280)/A(260) ratio can also result from contamination with UDP-linked murein precursors derived from bacterial cell wall metabolism. Although these precursors are small molecules of molecular weight 1000-1200, they comigrate in gel filtration with recombinant human FKBP (MW 11,820). This gel filtration behavior, which is distinct from that of unmodified mononucleotides, does not reflect binding interactions with FKBP, but is an intrinsic property of these precursors. Therefore, these molecules would be expected to copurify with other low-molecular-weight proteins, especially in the abbreviated purification protocols made possible by freeze-thaw release of recombinant proteins from E. coli (Johnson, B. H., and Hecht, M. H. (1994) BioTechnology 12, 1357-1360). Several alternative strategies are discussed for integrating these findings into the design of improved purification procedures for low-molecular-weight recombinant proteins.     

Structure and variability of mammalian peroxisomal membrane proteins.

Peroxisomal membrane proteins (PMPs) from the Swiss-Webster mouse are analyzed and compared to those of rats and humans using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. A purification procedure for fresh mouse, rat, or human biopsy liver which enriches peroxisomal/mitochondrial marker enzyme ratios over 100-fold is characterized. When analyzed by SDS-PAGE, membranes of purified liver peroxisomes are shown to contain the same complement of 145-, 70-, 55-, 36-, and 22-kDa PMPs in rats, mice, and humans. A rabbit polyclonal antibody raised against mouse peroxisomal membranes demonstrates immunoreactivity to 145- and 70-kDa proteins in fresh liver homogenates from all three species and in control or Zellweger syndrome fibroblasts from humans. Human autopsy or placental tissues which were refrigerated before analysis exhibited 105-, 55-, and 36-kDa peptides which may be derived from the 145- and 70-kDa peptides. Such conversions, if related to degradation, may explain difficulties in purifying peroxisomes from human autopsy specimens. Variable amounts of the 55-kDa peptide also occurred in mouse adrenal and lung, and the conversion of higher to lower molecular weight PMPs could not be demonstrated by in vitro incubation of mouse liver. Further definition of the structure and variability of mammalian PMPs should be helpful in understanding polyenzymopathies such as Zellweger syndrome.