Contributions of cytoplasmic free and membrane-bound ribosomes to the synthesis of NADPH-adrenodoxin reductase and adrenodoxin of bovine adrenal cortex mitochondria

Cytoplasmic free and membrane-bound ribosomes were isolated from bovine adrenal cortex, and characterized. Contributions of free and bound ribosomes to the synthesis of NADPH-adrenodoxin reductase (AdR) and adrenodoxin (Ad) were determined by examining the presence of their nascent peptides on isolated ribosomes. Nascent peptides were released from the ribosomes by [3H]puromycin in a high salt buffer in the presence of a detergent, and the nascent peptides of AdR and Ad were separately isolated by immunoprecipitation using antibodies. AdR nascent peptides were associated with free and loosely-bound ribosomes, whereas Ad nascent peptides were associated with free, loosely-bound and tightly-bound ribosomes. Smaller nascent peptides of AdR were carried by free ribosomes, whereas larger nascent peptides were preferentially carried by loosely-bound ribosomes. In the case of Ad, smaller nascent peptides were more abundant in free ribosomes than in bound ribosomes. The nascent peptides of Ad were released from bound ribosomes of rough microsomes to the aqueous milieu by puromycin treatment, suggesting the release of completed Ad peptides into the cytoplasm in cells.     

Diversity of wheat anti-microbial peptides

From seeds of Triticum kiharae Dorof. et Migusch., 24 novel anti-microbial peptides were isolated and characterized by a combination of three-step HPLC (affinity, size-exclusion and reversed-phase) with matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry and Edman degradation. Based on sequence similarity and cysteine motifs, partially sequenced peptides were assigned to 7 families: defensins, thionins, lipid-transfer proteins, hevein-like peptides, knottin-like peptides, glycine-rich peptides, and MBP-1 homologs. A novel subfamily of defensins consisting of 6 peptides and a new family of glycine-rich (8 peptides with different repeat motifs) were identified. Three 6-cysteine knottin-like peptides represented by N- and C-terminally truncated variants revealed no sequence homology to any known plant anti-microbial peptides. A new 8-cysteine hevein-like peptide and three 4-cysteine peptides homologous to MBP-1 from maize were isolated. This is the first communication on the occurrence of nearly all families of plant anti-microbial peptides in a single species.     

A Convenient Synthesis of 4-Hydroxy-3-methyl-2-(2-propynyl)-cyclopent-2-enone,an Alcohol Moiety of a Synthetic Pyrethroid Having Strong Killing and Knockdown Activity

We examined the primary structure of the α-amylase produced by Bacillus subtilis var. amylosacchariticus by attempting to isolate tryptic peptides of the enzyme. By solubilization and precipitation in buffers, the peptides were first fractionated into three. The main fraction was fractionated by ion-exchange chromatography. Twenty-seven peptides were generated from this fraction. The fraction insoluble at neutral pH was fractionated by SP-Sephadex C-25. From this fraction three peptides were obtained. The other fraction insoluble at acidic pH was fractionated by Bio-Gel P-60. Four peptides were isolated from this fraction. In total, thirty-four peptides were generated from the tryptic digest of the α-amylase. The amino acid sequences of twenty-one out of thirty-four peptides were completely determined, while those of the other thirteen peptides were partially determined. The peptides derived from the N- and C-terminal ends of the α-amylase were identified.     

Kinins in humans

The kinin peptide system in humans is complex. Whereas plasma kallikrein generates bradykinin peptides, glandular kallikrein generates kallidin peptides. Moreover, a proportion of kinin peptides is hydroxylated on proline(3) of the bradykinin sequence. We established HPLC-based radioimmunoassays for nonhydroxylated and hydroxylated bradykinin and kallidin peptides and their metabolites in blood and urine. Both nonhydroxylated and hydroxylated bradykinin and kallidin peptides were identified in human blood and urine, although the levels in blood were often below the assay detection limit. Whereas kallidin peptides were more abundant than bradykinin peptides in urine, bradykinin peptides were more abundant in blood. Bradykinin and kallidin peptide levels were higher in venous than arterial blood. Angiotensin-converting enzyme inhibition increased blood levels of bradykinin, but not kallidin, peptides. Reactive hyperemia had no effect on antecubital venous levels of bradykinin or kallidin peptide levels. These studies demonstrate differential regulation of the bradykinin and kallidin peptide systems, and indicate that blood levels of bradykinin peptides are more responsive to angiotensin-converting enzyme inhibition than blood levels of kallidin peptides.     

多肽的α螺旋结构对多肽与钙调蛋白亲合力的影响

本文设计并采用固相法合成了4种钙调蛋白可结合多肽,这些多肽分成两组,每一组中两个多肽的碱性和疏水性相近,但形成α螺旋结构的倾向(预测)不同。研究了这些多肽与钙调蛋白的相互作用,在Ca~(2+)存在下,这些多肽与丹磺酰钙调蛋白结合,使丹磺酰钙调蛋白的荧光光谱发生显著变化,测定了多肽与钙调蛋白所形成的复合物的解离常数。结果表明,预测形成α螺旋结构倾向较大的多肽与钙调蛋白的亲合力也较大。     

Bovine hemoglobin: an attractive source of antibacterial peptides

A peptic hemoglobin hydrolysate was fractioned by a semi-preparative reversed-phase HPLC and some fractions have an antibacterial activity against four bacteria strains: Micrococcus luteus A270, Listeria innocua, Escherichia coli and Salmonella enteritidis. These fractions were analyzed by ESI/MS and ESI/MS/MS, in order to characterize the peptides in these fractions. Each fraction contains at least three peptides and some fractions contain five peptides. All these fractions were purified several times by HPLC to obtain pure peptides. Thirty antibacterial peptides were identified. From the isolated antibacterial peptides, 24 peptides were derived from the chains of hemoglobin and 6 peptides were derived from the β chains of hemoglobin. The lowest concentration of these peptides (minimum inhibitory concentration (MIC)) necessary to completely inhibit the growth of four bacteria strain was determined. The cell population of all of the tested bacteria species decreased by at least 97% after a 24-h incubation with any of the peptides at the minimum inhibitory concentration.     

The primary structure of actin from rabbit skeletal muscle. Three cyanogen bromide peptides that are insoluble at neutral pH.

Three of the 17 peptides produced when actin is treated with cyanogen bromide are sparingly soluble at pH values near neutrality. They were separated from more soluble peptides at pH 6.0 on a column of Sephadex G-10. The soluble peptides were excluded from the gel and emerged at the void volume, while the insoluble peptides were "washed off" by the formic acid in which the sample was applied. The three insoluble peptides were sequenced as a group by studying peptides generated by tryptic and chymotryptic digestion of the mixture, and peptic digestion of the partially resolved peptides. The three peptides are: CB-15 (residues 133 to 176), CB-16 (residues 325 to 354), and CB-17 (residues 191 to 227).     

Solid phase synthesis of hydrophobic difficult sequence peptides on BDDMA-PS support.

This article illustrates the successful and efficient solid phase assembly of hydrophobic difficult sequence peptides following both t-Boc and Fmoc chemistry. The peptides were synthesized on an optimized 1,4-butanediol dimethacrylate-crosslinked polystyrene support (BDDMA-PS). Four difficult sequence test peptides, VAVAG, VIVIG, QVGQVELG and VQAAIDYING, were synthesized in relatively good yield and purity without any aggregation problems. The peptides were assembled on chloromethylated and 4-hydroxymethylphenoxymethyl (HMP) BDDMA-PS resins. The peptides were fabricated using Boc amino acid 1-hydroxybenzotriazolyl and Fmoc amino acid pentafluorophenyl active esters in coupling reactions. The peptides after synthesis were cleaved from the polymeric support by exposing the peptidyl resin to 90% trifluroacetic acid/5% thioanisole/5% EDT mixture. The HPLC and MALDI TOF MS studies of the peptides revealed the high homogeneity of the synthesized peptides. Chloromethylated resin having a functional group loading of 1.14 mmol Cl/g was used for the synthesis. The yield and homogeneity of these peptides synthesized using the new support were high when compared with the conventional DVB-PS resin.     

HLA-DR-presented peptide repertoires derived from human monocyte-derived dendritic cells pulsed with blood coagulation factor VIII

Activation of T-helper cells is dependent upon the appropriate presentation of antigen-derived peptides on MHC class II molecules expressed on antigen presenting cells. In the current study we explored the repertoire of peptides presented on MHC class II molecules on human monocyte derived dendritic cells (moDCs) from four HLA-typed healthy donors. MHC class II-bound peptides could be routinely recovered from small cultures containing 5 × 10(6) cells. A fraction of the identified peptides were derived from proteins localized in the plasma membrane, endosomes, and lysosomes, but the majority of peptides that were presented on MHC class II originate from other organelles. Subsequently, we studied the antigen-specific peptide repertoire after endocytosis of a soluble antigen. Blood coagulation factor VIII (FVIII) was chosen as the antigen since our current knowledge on MHC class II presented peptides derived from this immunogenic therapeutic protein is limited. Analysis of the total repertoire of MHC class II-associated peptides revealed that per individual sample 20-50 FVIII-derived peptides were presented on FVIII-pulsed moDCs. Repertoires of FVIII-derived peptides eluted from moDCs derived from a panel of four HLA typed donors revealed that some MHC class II-presented FVIII peptides were presented by multiple donors, whereas the presentation of other FVIII peptides was donor-specific. In total 32 different core peptides were presented on FVIII-pulsed moDCs from four HLA-typed donors. Together our findings provide an unbiased approach to identify peptides that are presented by MHC class II on antigen-loaded moDCs from individual donors.     

蛙属Rana活性肽研究新进展

本文对近年从蛙属Ｒａｎａ提取分离的活性肽作了综述,对一些活性肽如排钠利尿肽,速激肽,抗菌肽,激素肽等作了较详细的介绍。