Fluorine concentrations in bone biopsy samples determined by proton-induced gamma-ray emission and cyclic neutron activation

Fluorine concentrations in bone biopsy samples taken from the iliac crest of subjects, divided into four groups depending on the length of dialysis treatment, and aluminium levels in blood and bone pathology, in terms of osteoporosis, were determined by two instrumental methods. Proton-induced gamma-ray emission (PIGE), making use of the resonance reaction of¹⁹F(p,αγ)¹⁶O at 872 keV, and cyclic neutron activation analysis (CNAA), using the¹⁹F(n,γ)²⁰F reaction in a reactor irradiation facility, were employed. Rutherford backscattering (RBS) was used to calculate the volume, and, hence, mass of the sample excited in PIGE by determining the major element composition of the samples in order to express results in terms of concentration. From this preliminary investigation, a relationship is suggested between fluorine concentrations in bone and aluminium levels in the system.

     

On the Permeation by Dioxygen of the Cofactor‐Independent Unusual Oxygenase RhCC,in Complex with Substrate 4‐Hydroxyphenylenolpyruvate. A Molecular Dynamics Investigation

This work deals with a trimeric bacterial protein, RhCC, which, although belonging to the tautomerase superfamily, shows oxygenase activity. A model of the complex from RhCC and substrate 4‐hydroxyphenylenolpyruvate (4HPP), fitting the observation of extra electron densities from X‐ray diffraction of the crystal, could be built by autodocking. When subjected to molecular dynamics (MD) aided by an external random force applied to a O₂ molecule placed above 4HPP, this model evolved with O₂ egressing toward the bulk solvent from two nearly opposite gates. These were located between the nearly parallel helices 75 – 91 and 15 – 33 of either chain C (gate SE) or chain B (gate FL). Alternatively, with four O₂ molecules in the bulk solvent, unbiased MD led to O₂ entering the protein from gate SE and getting to 4HPP, while forming a stabilizing salt bridge between the 4HPP carboxylate and P1.C ⁺NH₂, thus providing scientific ground for a refined model of the complex.     

Effects of EDTA treatment upon the protein subunit composition and mechanical properties of mammalian single skeletal muscle fibers

Considerable interest has been focused on the role of myosin light chain LC(2) in the contraction of vertebrate striated muscle. A study was undertaken to further our investigations (Moss, R.L., G.G. Giulian, and M.L. Greaser, 1981, J. Biol. Chem., 257:8588-8591) of the effects of LC(2) removal upon contraction in skinned fibers from rabbit psoas muscles. Isometric tension and maximum velocity of shortening, V(max), were measured in fiber segments prior to LC(2) removal. The segments were then bathed at 30 degrees C for up to 240 min in a buffer solution containing 20 mM EDTA in order to extract up to 60 percent of the LC(2). Troponin C (TnC) was also partially removed by this procedure. Mechanical measurements were done following the EDTA extraction and the readditions of first TnC and then LC(2) to the segments. The protein subunit compositions of the same fiber segments were determined following each of these procedures by SDS PAGE of small pieces of the fiber. V(max) was found to decrease as the LC(2) content of the fiber segments was reduced by increasing the duration of extraction. EDTA treatment also resulted in substantial reductions in tension due mainly to the loss of TnC, though smaller reductions due to the extraction of LC(2) were also observed. Reversal of the order of recombination of LC(2) and TnC indicated that the reduction in V(max) following EDTA treatment was a specific effect of LC(2) removal. These results strongly suggest that LC(2) may have roles in determining the kinetics and extent of interaction between myosin and actin.     

Occurrence of 5''-Deoxy-5''-Methylthioadenosine Phosphorylase in the Mammalian CNS: Distribution and Kinetic Studies on the Rat Brain Enzyme

5'-Deoxy-5'-methylthioadenosine (MTA) phosphorylase catalyzes the cleavage of MTA, a secondary product of polyamine biosynthesis, to 5-methylthioribose-1-phosphate and adenine. The occurrence and the general properties of the enzyme were studied in mammalian brain with the following results. (1) Cerebral tissues contained levels of MTA phosphorylase that were comparable to those occurring in other mammalian tissues. (2) Interspecies differences in the enzyme distribution were quite limited, with the highest specific activity values observed in pig brain. Moreover, the enzyme seemed to be generally more concentrated in the cerebellar fractions. (3) Rat brain MTA phosphorylase was highly localized in the cellular soluble fraction. In the first days of rat life, its specific activity in the whole brain was observed to decline significantly from a value of 17.6 units/mg at 1-5 days of age to 13.7 units/mg at 6-10 days of age, remaining then fairly constant up to maturity. (4) Kinetic studies performed with the soluble enzyme extracted from rat brain showed: a pH optimum of 7.4; a Km value for MTA of about 10 microM; an inhibitory effect of the MTA analog 5'-deoxy-5'-isobutylthioadenosine; and a remarkable resistance of the enzyme to heat treatment.     

From the Sequence to the Conformation of the Unabridged Transmembrane Domains TM1 and TM2 of the cASIC1a Ion Channel – A Parallel Tempering Approach

This work was devised to unravel, along replica‐exchange molecular‐dynamics (REMD) simulations, the conformation in solution of the TM1 and TM2 transmembrane domains of the homotrimeric cASIC1a ion channel. This includes the head of TM1 and tail of TM2 that had previously defied X‐ray diffraction analysis in the crystal. The structure of the open‐channel complex of cASIC1a with psalmotoxin 1 (PcTx1) was chosen here as a basis, although, to make the simulations affordable, the procedure was limited to the missing portions, including a few adjacent α‐helical turns. The latter were held fixed during the simulations. Reassembling the whole subunit, by superimposition of the fixed portions, resulted in diving of both TM1 and TM2 as continuous α‐helices into the cytoplasm. At completion of this work, it appeared, from similar X‐ray diffraction studies, that TM2 for both the complex of cASIC1a with the coral snake MitTx toxin, and the isolated desensitized ion channel, is discontinuous, with the triad G443‐A444‐S445 taking an extended, belt‐like conformation. In this way, a filter ring against hydrated ions is formed by G443 in the trimer. Our REMD examination of this complex revealed a strong resistance by G443, and only that residue, to take dihedral‐angle values compatible with an α‐helical conformation. This suggests that the flexibility of glycine alone does not explain formation of the extended, belt‐like conformation of the triad G443‐A444‐S445. This also requires cooperation in the trimer.