Adenosine-5′-d: rotational conformation of the 5′-carbinol group

A synthesis of adenosine-5′-d (4), and its p.m.r. spectral characteristics, are described. The presence of deuterium in 4 gives rise to a 2:1 mixture of R and S configurations at C-5, thereby permitting specific assignments for the resonances of the residual 5′-protons. From the observed spin-spin coupling between the latter and H-4′, and estimate has been made of the rotamer population of the exocyclic 5′-carbinol group. It is shown that the gauche-gauche rotamer is preponderant (≈70%) and the gauche-trans one of minor importance (≈20%) in aqueous solution, which contrasts markedly with the preference for the latter rotamer exhibited by adenosine in the solid state.     

Postmortem Changes in Rat Brain: Studies on Membrane-Bound Enzymes and Receptors

The relationship between the stability of potential neurochemical markers and autolysis time was studied at 4 degrees C and 25 degrees C using postmortem brain samples from two rat strains. In general, qualitatively similar results were obtained with either N/Nih or Sprague-Dawley rats; however, quantitative differences were often observed, particularly in regard to benzodiazepine receptor changes. For every enzyme activity or binding property examined, no significant change was found when brains were kept at 4 degrees C for up to 72 h prior to freezing at -70 degrees C. Na,K-ATPase and low-affinity Ca-ATPase activities were also stable in brains kept at 25 degrees C for up to 72 h. Mg-ATPase activity was reduced in brains kept at 25 degrees C for 24 and 48 h. [3H]Guanidinoethylmercaptosuccinic acid [( 3H]GEMSA) binding to enkephalin convertase in the cytosol was not significantly changed in brains kept at 25 degrees C; however, a small increase was seen for [3H]GEMSA binding to the membrane fraction at 24, but not 48 and 72 h postmortem. [3H]Quinuclidinyl benzilate [( 3H]QNB) binding to muscarinic cholinergic receptors decreased in brains kept at 25 degrees C for 72 h. Opioid receptor binding also decreased in brains kept at 25 degrees C. Using [3H]2-D-alanine-5-D-leucine enkephalin to label delta opioid receptors, a statistically significant decrease in binding was observed as early as 6 h postmortem, and was completely abolished after 72 h at 25 degrees C. In contrast, [3H]naloxone binding was unchanged after 24 h at 25 degrees C, but was decreased after 48 and 72 h.(ABSTRACT TRUNCATED AT 250 WORDS)     

Novobiocin-resistance sequences from the novobiocin-producing strain Streptomyces niveus

Two distinct DNA sequences expressing novobiocin resistance in Streptomyces lividans were cloned from the novobiocin-producing species Streptomyces niveus. Clone pGL101 (5kb) conferred resistance to 50 micrograms ml-1 novobiocin, whereas clones pGL102 and pGL103, which carry the same 6.5kb insert but in opposite orientations, expressed resistance to 150 micrograms ml-1. The cloned inserts from pGL101 and pGL103 failed to hybridize with each other or with the cloned novobiocin-resistant gyrB sequence from Streptomyces sphaeroides. Both probes hybridized strongly with DNA from the novobiocin-producing species S. niveus and S. sphaeroides but no hybridization (pGL103) or very weak hybridization (pGL101) was detected with DNA from the non-producing species S. lividans, Streptomyces griseus and Streptomyces antibioticus. S. niveus contains at least three novobiocin-resistance determinants with the pGL101 and pGL103 cloned sequences specific for novobiocin-producing strains of Streptomyces.     

Restriction pattern and polypeptide homology among plasmid-borne mercury resistance determinants

The structural and functional properties of mercury resistance determinants cloned from a series of independently isolated conjugative plasmids were compared with those of the prototype HgR determinants from Tn501 and plasmid R100 (containing Tn21). Restriction endonuclease mapping classified the HgR determinants into at least three different but related structural groups which are distantly related to those from Tn501 and R100. These relationships were confirmed by the functional analysis of sub-clones and gamma delta insertion mutations and from the polypeptides specified by the cloned HgR determinants. Each mercury resistance clone synthesized polypeptides equivalent in size to the merA, merT, and merP gene products. However, those for merA and merT showed considerable size variation. No polypeptide equivalent to merD or merC of R100 was detected.     

Phage T1-mediated transduction of a plasmid containing the T1 pac site

The T1 pac site has been cloned into a plasmid vector. This recombinant plasmid was tested for T1-mediated transduction efficiency in comparison with a plasmid containing the phage lambda T1-pac-like site esp-lambda, plasmids containing T1 sequences other than the pac site, and plasmids containing neither T1 sequences nor known pac sites. The data obtained indicate that there are at least two distinct mechanisms of T1-mediated plasmid transduction. One requires the presence of any T1 sequence on the plasmid and probably takes place via cointegrate formation with the homologous region of an infecting T1 genome. The other is specifically dependent on the presence of a pac site on the plasmid. Plasmids are packaged as head-to-tail multimers that have one heterogeneous molecular end and the other terminated at pac, and the direction of packaging with respect to the pac site is the same for plasmids as for T1. Possible roles of pac in plasmid packaging and their implications with regard to the packaging of phage DNA are discussed.     

Mutant of Salmonella typhimurium LT2 deficient in DNA adenine methylation.

A mutant of Salmonella typhimurium LT2 deficient in methylation of the adenine residues in the sequence 5'-GATC-3' was isolated. The mutation (dam-1) was linked to the cysG locus, and the properties of the mutant were similar to those of Escherichia coli dam mutants. Reversion of the hisC3076 frameshift marker by 9-aminoacridine was substantially enhanced by the dam-1 mutation, implying a direct role for adenine methylation in the prevention of frameshift mutation induction.     

Stability of neuronal and glial marker enzymes in post-mortem rat brain

The enzymatic activities in post-mortem rat brain kept at 4°C and at 25°C were determined for a number of enzymes localized in specific cell types in the central nervous system. Choline acetyltransferase (CAT), glycerol-3-phosphate dehydrogenase (GPDH), glutamine synthetase (GS), lactate dehydrogenase (LDH) and 2,3-cyclic nucleotide phosphohydrolase (CNPase) were found to be very stable at both 4°C and 25°C with only slight, if any, losses of activity being seen even at periods as long as 72 hr. Glutamic acid decarboxylase (GAD) activity was less stable than that of the other enzymes. In brains kept at 4°C GAD activity was stable out to 24 hr after which it began to decline rapidly to 65% of control at 72 hr. In brains kept at 25°C, GAD activity was stable for 6–8 hr and then began to steadily decline to 58% of control at 24 hr and 29% of control at 72 hr. Assuming that these enzymes have similar stabilities in post-mortem human brain, the effect of post-mortem delay in processing tissues may be of lesser significance than other factors with regard to the measured enzyme activities in human brain samples.