"In vivo" (35S)-methionine interaction with rat-liver DNA and the effect of chemical carcinogens

1. After intraperitoneal administration of (35S)methionine (25 mg, 1.6 mCi/kg), detectable amount of radioactivity resulted associated to rat-liver DNA: the interaction reached the maximum value (about 18 pmol/mumol DNA P) by 2 h after administration of radioactive aminoacid. 2. The (35S)-binding was inhibited by the hepatocarcinogenic ethionine and dimethylnitrosamine, and was stimulated by the non-hepatocarcinogenic methylnitrosourea. 3. Hplc analysis of (35S)DNA enzymic digest evidenced two radioactive compounds, the UV behaviour of which is reported.     

Differential D-glucose requirements of the general amino-acid permease and protein synthesis in Saccharomyces cerevisiae var. ellipsoideus

The dependence of the general aminoacid permease and protein synthesis on the availability of D-glucose as energy source was studied. Stimulation by the sugar was immediate once added to the cell suspensions and seems to be mediated by energy derived directly from glycolysis. The general aminoacid permease was saturated linearly with D-glucose whereas protein synthesis was saturated sigmoidealy requiring much higher concentration of the sugar than the general aminoacid permease.     

落叶果树花粉中的氨基酸含量及其评价

<正> 蜜源植物蜂花粉,在营养学上被称作“蛋白质和氨基酸的浓缩体”。落叶果树中的苹果花粉,也被确认为营养价值极好级别的植物花粉之一。但是,对于落叶果树花粉中的氨基酸含量,树种、品种间的区别,迄今国内外尚少系统研究和报道。为了认识、评价和选择利用落叶果树的花粉资源,于1987～1989年间,对10个落叶果树树种、82个品种花粉中的氨基酸含量,进行了系统的测定、分析。现将结果整理报告于下。     

Effect of different oxygen pressure levels on the transport of alpha-aminoisobutyric acid in myocardial cell cultures

The activity of aminoacid transport, as measured by alpha-aminoisobutyrate uptake, has been studied in confluent myocardial cell cultures exposed to different oxygen tensions. The results obtained indicate that the rate of cellular uptake and accumulation of the inert aminoacid increase with time as the fraction of oxygen is reduced. When alpha-aminoisobutyrate was added in presence of all other aminoacids of the medium, the effect of oxygen was also evident, suggesting a mechanism which overcomes the competitive action of the other aminoacids assigned to the same transport system of alpha-aminoisobutyrate (A system). The modulation of aminoacid transport activity may represent one of the possible mechanisms by which environmental oxygen affect the rate of cellular protein synthesis.     

A rapid and sensitive method for the estimation of individual tRNA pools

A rapid and sensitive method is described to quantitatively compare tRNA pools for individual aminoacids in a single experiment. The procedure comprises of: charging of total tRNA with a mixture of radiolabeled aminoacids, deacylation of the esterified tRNA with a volatile base and the recovery of the labeled aminoacid, derivatisation of the aminoacid with phenylisothiocyanate after mixing with excess of nonradioactive aminoacids, baseline separation of the phenylthiocarbamyl aminoacids by reverse phase high performance liquid chromatography monitored by A254nm and quantitation of the radioactivity in individual aminoacid peaks. The radioactivity in the aminoacid peak corresponds to the quantity of the aminoacylated tRNA. The method has been successfully applied to quantitate the individual tRNA pools in the developing silk glands of Bombyx mori, a functionally adapted tissue which undergoes considerable variations in tRNA content.     

Chemical analysis of cryoglobulins observed in a case of essential IgG-IgA cryoglobulinaemia

In a case of essential mixed (IgG-IgA) cryoglobulinaemia the carbohydrate and aminoacid composition of cryoglobulins was chemically analyzed. A normal aminoacid and sialic acid composition was observed, whereas the glucide percentage was augmented with respect to the normal range for the two classes of Ig.     

Addition of amino acids and oligopeptides to the diet of piglets. I. Effects on productive performance and development of intestinal microbial flora

The effects in connection with the addition of small quantities of aminoacid and oligopeptide pools to balanced diets were studied on 36 male piglets, divided into three groups (1: control; 2: aminoacid pool; 3: aminoacid and oligopeptide pool). Piglets receiving the aminoacid pool (group 2) exhibited better performances: an increased weight gain resulting in higher feed efficiency and an increased overall microbial count with a predominance of the useful population.     

Influence of hydrophobic amino acid residues on the esterification of dipeptides by papain

Summary Esterification of dipeptide was performed with papain immobilized on XAD-7 with methylene chloride as the organic solvent. The esterification of a peptide substrate Box-X-Y-OH is promoted by the presence of a hydrophobic aminoacid residue (e.g., Phe, Leu) in the X (P2) position. Good yields were observed for example with Boc-Phe-Glu-OH and Boc-Ala-Glu-OH. With a hydrophilic aminoacid residue in the X position no esterification of the dipeptide takes place. Instead hydrolysis of the peptide bond occurs. This is also the case with glycine as the aminoacid; for instance Boc-Leu-Gly-OH gives a 80% yield of esterification whereas Boc-Gly-Leu-OH gives only the aminoester Boc-Gly-OR.     

Molecular structure of the immunity gene and immunity protein of the bacteriocinogenic plasmid Clo DF13.

The nucleotide sequence of the Clo DF13 DNA region comprising the immunity gene has been determined. We also elucidated the aminoacid sequence of the 40 N-terminal and 7 C-terminal aminoacids of the purified immunity protein. From analysis of the data obtained we were able to locate the immunity gene between 11.7 and 14.5% on the Clo DF13 map, and to determine the complete aminoacid sequence of the immunity protein. It was observed that the Clo DF13 immunity gene encodes an 85 aminoacid protein and is transcribed in the same direction as the cloacin gene. These experimental data support our model, presented elsewhere, which implicates that the cloacin and immunity genes of Clo DF13 are coordinately transcribed from the cloacin promoter. We also present DNA sequence data indicating that an extra ribosome binding site precedes the immunity gene on the polycistronic mRNA. This ribosome binding site might explain the fact that in cloacinogenic cells more immunity protein than cloacin is synthesized. The comparison of the complete aminoacid sequence of the Clo DF13 immunity protein, with the aminoacid sequence data of the purified, comparable Col E3 immunity protein revealed that both proteins have extensive homologies in primary and secondary structure, although they are exchangeable only to a low extent in vivo and in vitro. It was also observed that a lysine residue was modified in immunity protein isolated from excreted bacteriocin complexes.     

BS01菌株降解石蜡产生物表面活性剂的研究

从辽河油田排气池水样中分离出一株能降解石蜡产生物表面活性剂的杆菌BS01菌株。根据传统分类和系列鉴定,该菌为铜绿假单胞菌。用通过正交试验得到的最佳培养基进行摇瓶培养,6d后,培养液的表面张力降低了12mN/m,pH降低了2～3个单位。经氨基酸自动分析仪和气象色谱测定,培养液中含有多种氨基酸和少量脂肪酸。培养液用正已烷多次萃取后,有机相经紫外分光光度计测定,石蜡降解率为69％。     

Aminoacid composition and hydrophobicity index of mitochondrial polypeptides

The aminoacid composition of protein stained bands in polyacrylamide gels, after electrophoresis of proteins from inner mitochondrial membranes, was investigated hydrolyzing directly the gel slices. The Hydrophobicity Index of 18 prominent polypeptide bands was calculated after their aminoacid analysis. The polypeptides less related to the membrane have low hydrophobicity as inferred from their Hydrophobicity Indexes.     

Two forms of methionyl-transfer RNA synthetase from Mycobacterium smegmatis

Two methionyl-transfer RNA synthetases (A and B forms) have been isolated from $\text{Mycobacterium smegmatis}$. The homogeneous preparations of the enzymes showed 1500 fold increase in specific activity in aminoacylation of methionine specific tRNA. The A and B forms differed in their specificity of aminoacylation of tRNA_m^Met and tRNA_f^Met; enzyme B exhibited much higher specificity for tRNA_f^Met. The molecular activities of A and B enzymes for aminoacid and tRNA were identical. The turnover number for aminoacid was 27 fold greater than that for tRNA, while the Km values for tRNA were lower by a factor of 10⁶ as compared to the aminoacid. Both the enzymes catalysed ATP-pyrophosphate exchange reaction to the same extent.     

Hb Sinai,a new α chain mutant a 47 His

Summary A new chain mutant Hb-Sinai ₄₇ ^His is described. The aminoacid composition of all tryptic peptides has been determined, with the exception of the insoluble core. In the fingerprint peptide T 6 which normally migrates between T 3 and T 1, moves now between T 3 and T 7. The aminoacid composition of peptide T 6 indicates a change in the aminoacid composition from Asp- to His⁺ in position 47.This work was supported by Grant No. GM 13714 U.S.P.H.S.     

Isolation and characterization of Post Gamma globulin in mouse

A protein of about 13,000 daltons was isolated from mouse CBA urine after inducing a tubular renal dysfunction. This protein was demonstrated similar to human Post Gamma globulin by electrophoresis, aminoacid content and immunochemical criteria.     

The imbalance of brain large-chain aminoacid availability in amyotrophic lateral sclerosis patients treated with high doses of branched-chain aminoacids

Following the report of an increased mortality among patients with amyotrophic lateral sclerosis given high daily doses of branched-chain aminoacids, we assessed the plasma concentrations of large neutral aminoacids and glutamic acid and the large neutral aminoacid brain influx in 24 amyotrophic lateral sclerosis patients receiving placebo or branched-chain aminoacids ( -leucine 12 g, -isoleucine 6 g, -valine 6 g daily), in 15 untreated amyotrophic lateral sclerosis patients and in 15 healthy volunteers. The branched-chain aminoacid plasma concentrations increased three- to six-fold in the treated group compared to the patients receiving placebo or no treatment and to the healthy controls. Plasma glutamic acid concentrations in healthy volunteers were 51.59±7.53 nmol/ml while in the amyotrophic lateral sclerosis patients receiving no treatment, placebo or branched-chain aminoacids were 92.33±12.15 nmol/ml, 91.21±15.86 nmol/ml and 95.08±17.96 nmol/ml respectively. The glutamic acid concentration was significantly higher (P<0.01) in amyotrophic lateral sclerosis patients than in healthy individuals. Plasma phenylalanine and tyrosine were lower in the amyotrophic lateral sclerosis patients than in healthy controls, regardless of treatment, whereas tryptophan levels were not significantly different. The branched-chain aminoacid brain influx of the treated group was 110-140% of that measured in the patients receiving placebo and in the healthy controls. The aromatic aminoacid brain influx was lower in the treated group than in the placebo group or healthy controls. An impairment of brain large neutral aminoacid availability might possible contribute to enhancing the progression of symptoms in patients with amyotrophic lateral sclerosis.     

Visualization and quantification of central 5-HT1A receptors with specific antibodies

Polyclonal antibodies were raised by the repeated injection of rabbits with synthetic peptides corresponding to selective portions (peptide 1: aminoacid residues 12–23, and peptide 2: aminoacid residues 243–268) of the aminoacid sequence of the rat 5-HT_1A receptor. Both antisera allowed the immunoprecipitation of 5-HT_1A receptors but not of other 5-HT receptor types and adrenergic receptors solubilized from rat hippocampal membranes. Immunoblots demonstrated that a single protein of 63 kDa, corresponding to the molecular weight of the rat 5-HT_1A receptor binding subunit, was recognized by each antiserum. Immunoautoradiographic labelling of rat brain sections with the anti-peptide 2-antiserum exhibited the same regional distribution as 5-HT_1A sites labelled by selective radioligands such as [³H]8-OH-DPAT and [¹²⁵I]BH-8-MeO-N-PAT. However regional differences apparently existed between the respective intensity of labelling by the agonist radioligands and the antiserum, which might be explained by variations in the degree of coupling of 5-HT_1A receptor binding subunits with G proteins from one brain area to another.     

Computational protein design with a generalized Born solvent model: application to Asparaginyl-tRNA synthetase

Computational Protein Design (CPD) is a promising method for high throughput protein and ligand mutagenesis. Recently, we developed a CPD method that used a polar-hydrogen energy function for protein interactions and a Coulomb/Accessible Surface Area (CASA) model for solvent effects. We applied this method to engineer aspartyl-adenylate (AspAMP) specificity into Asparaginyl-tRNA synthetase (AsnRS), whose substrate is asparaginyl-adenylate (AsnAMP). Here, we implement a more accurate function, with an all-atom energy for protein interactions and a residue-pairwise generalized Born model for solvent effects. As a first test, we compute aminoacid affinities for several point mutants of Aspartyl-tRNA synthetase (AspRS) and Tyrosyl-tRNA synthetase and stability changes for three helical peptides and compare with experiment. As a second test, we readdress the problem of AsnRS aminoacid engineering. We compare three design criteria, which optimize the folding free-energy, the absolute AspAMP affinity, and the relative (AspAMP-AsnAMP) affinity. The sequences and conformations are improved with respect to our previous, polar-hydrogen/CASA study: For several designed complexes, the AspAMP carboxylate forms three interactions with a conserved arginine and a designed lysine, as in the active site of the AspRS:AspAMP complex. The conformations and interactions are well maintained in molecular dynamics simulations and the sequences have an inverted specificity, favoring AspAMP over AsnAMP. The method is not fully successful, since experimental measurements with the seven most promising sequences show that they do not catalyze at a detectable level the adenylation of Asp (or Asn) with ATP. This may be due to weak AspAMP binding and/or disruption of transition-state stabilization.     

Partial purification and study of gamma-glutamyl transpeptidase from sheep cerebral capillaries]

Gamma-glutamyl transpeptidase was purified 53 times from Sheep brain cortex capillaries. On gel filtration it appears homogeneous with a M.W. = 350 000. The enzyme is likely a glycoprotein, the properties of which are close to hog kidney gamma-glutamyl transpeptidase ; gamma-glutamyl aminoacid formation is assayed electrophoretically. The results obtained using several aminoacids are in favour of the existence of different units, specific of each group of aminoacids ; together with the data from structural analogs, they support the hypothesis that gamma-glutamyl transpeptidase participates in aminoacid transport accross blood brain barrier.     

Characterization of enzymatic deficiencies of branched chain amino-acid catabolism in human fibroblasts by genetic complementation

Leucine and Isoleucine metabolism in cultured skin fibroblasts from patients with leucinosis, beta-Ketothiolase deficiency, propionic, methylmalonic and isovaleric acidemia, was compared with that in normal fibroblasts. A simple assay was developed using (U14C) Isoleucine and (U14C) Leucine as substrates. Radioactive incorporation into protein aminoacids were measured. The (U14C) branched chain aminoacid incorporation into proteins provides an estimation of the protein synthesis and the incorporation ratio (14C) Aspartate + (14C) Glutamate/(14C) branched chain aminoacid, measures the integrity of the metabolic pathway. Complementation tests permits to characterize the genetic defect. The incorporation ratio was significantly decreased in blocked pathways, namely in leucinosis and isovaleric acidemia in the presence of (U14C) Leucine and in Leucinosis, beta-Ketothiolase deficiency, propionic and methylmalonic acidemia in the presence of (U14C) Isoleucine. There was a significant restoration of activity in mutant strains with distinct genetic defects after polyethylene-glycol fusion. This assay provides a valuable tool to screen for enzymatic deficiencies of branched chain aminoacid catabolism.