摘要:

AbstractThe binding of the recombinant proteinase inhibitor eglincfrom the leechHirudo medicinalisto serine (pro)enzymes belonging to the chymotrypsin and subtilisin families has been investigated from the thermodynamic viewpoint, between pH 4.5 and 9.5 and from 10°C to 40°C. The affinity of eglincfor the serine (pro)enzymes considered shows the following trend: Leu‐proteinase [the leucine specific serine proteinase from spinach (Spinacia oleraceaL.) leaves]>human leucocyte elastase ⋍ human cathepsin G ⋍ subtilisin Carlsberg ⋍ bovine α‐chymotrypsin>bovine α‐chymotrypsinogen A ⋍ porcine pancreatic elastase ⋍ bovine β‐trypsin. The serine (pro)enzyme–inhibitor complex formation is an entropy‐driven process. On increasing the pH from 4.5 to 9.5, the affinity of eglincfor the serine (pro)enzymes considered increases thus reflecting the acid pKshift to the invariant hystidyl catalytic residue from ≈︁ 6.9 in the free serine proteinases and bovine α‐chymotrypsinogen A to ⋍ 5.1 in the serine (pro)enzyme–inhibitor complexes. Considering the known molecular models, the observed binding behaviour of eglincwas related to the inferred stereochemistry of the serine

ISSN:0952-3499    

DOI:10.1002/jmr.300040402

出版商:John Wiley&Sons, Ltd.    

年代:1991

数据来源: WILEY

摘要:

AbstractPhage G13 binds to the carbohydrate part of lipopolysaccharides from rough mutants ofSalmonellaandEscherichia colias the first event of infection. Equilibrium dialysis inhibition studies with native and synthetic trisaccharides as inhibitors suggested that phage G13 recognizes branched oligosaccharides having 6‐O‐α‐ or 7‐O‐α‐glycosyl groups with α‐Man(1→3) [α‐Man(1→6)]Man (Man[Man]Man) and α‐Glc(1→3)‐[α‐Hep(1→7)α‐Hep(1→3)α‐Hep(1→5) Kdo as the smallsest saccharides with inhibitory activity (Wollinet. al., 1989). Of four synthetic analogues to Man[Man]Man only Man(1→3)[α‐Gal(1→6)]α‐Man‐Ome (Man[Gal]‐Man) and α‐Glc(1→3)]α‐Hep(1→7)α‐Hep‐Ome (Glc[Hep]Hep) inhibited the binding of labelledE. coli.C core nonasaccharide ligand to G13 with activities which were 10‐ and 15‐fold lower than Man[Man]Man. The trisaccharides α‐Man(2→3)[α‐Glc(1→6)]α‐Man‐OMe (Man[Glc]Man) and α‐Man(1→3)[α‐Tal(1→6)]α‐Man‐OMe (Man[Tal]Man) showed no inhibition at concentrations 75‐fold higher than Man[Man]Man. Minimum energy conformation calculations of the saccharides using the GESA method showed that the 6‐O‐α‐Man group in Man[Man]Man and the 7‐O‐α‐Hep group in SL805 pentasaccharide expose their OH‐2 and OH‐3 groups in a similar way and these are postulated to be key structural features for binding activity. The importance of hydroxy groups at certain positions is implied from the fact that bothmanno‐ andgalacto‐isomers are active. We also conclude that the O6‐C6

ISSN:0952-3499    

DOI:10.1002/jmr.300040403

出版商:John Wiley&Sons, Ltd.    

年代:1991

数据来源: WILEY

摘要:

AbstractStudies using the T7 transcription system revealed that the discriminator base A73and the G20in the variable pocket play important roles in theEscherichia colialanine tRNA identity. The C60in the T‐loop, which is unique to alanine tRNA, was not found to be crucial for alanine identity. Anticodon replacement into the valine anticodon UAC did not decrease alanine charging activity, and no alanine charging activity was detected in the mutant valine tRNA possessing the alanine anticodon UG

ISSN:0952-3499    

DOI:10.1002/jmr.300040404

出版商:John Wiley&Sons, Ltd.    

年代:1991

数据来源: WILEY

摘要:

AbstractTo elucidate the mechanism of action of restriction endonucleasesMvaI andEcoRII a study was made of their interaction with a set of synthetic substrates in which the heterocyclic bases or the sugar–phosphate backbone had been modified; individual nucleotide residues had been removed or replaced with hydrocarbon bridges, and mismatched base pairs had been introduced. The groups of atoms in the heterocyclic bases and the phosphates in the recognition site that produce the most significant influence on the functioning of endonucleasesMvaI andEcoRII were discerned. Profound differences were found in the functioning of theMvaI andEcoRII neo‐schizomers. The catalytic activity ofEcoRII is significantly affected by any alteration in the recognition site structure and conformation, with a modification in one strand of the substrate causing the same decrease in the hydrolysis rate of both strands. EndonucleaseMvaI is tolerant to a number of structural abnormalities; the latter sometimes affect only hydrolysis of one strand of the recognition site. The enzyme can preferentially cleave one of the substrate strands. Mismatched base pairs retard and sometimes block the hydrolysis. The effect depends on the particular enzyme, mismatch and its locat

ISSN:0952-3499    

DOI:10.1002/jmr.300040405

出版商:John Wiley&Sons, Ltd.    

年代:1991

数据来源: WILEY

摘要:

AbstractWe have devised conditions whereby non‐tumorigenic, immunogenic cell variants of S49 mouse lymphoma were analyzed and separated from parental tumorigenic lymphoma cells. This was carried out using polyclonal antibodies (raised against the immunogenic variants) and immunomagnetic beads. The efficacy of the procedure depended on the amount of polyclonal antiserum, the immunobead to cell ratio, incubation time and the number of repetitions of the procedure. Experiments with mixed tumorigenic and non‐tumorigenic cells have resulted in an enrichment of up to 200‐fold of the non‐tumorigenic, immunogenic cells in the population. These findings indicate the potential use of this procedure (in conjunction with other approaches) to isolate from a population of tumorigenic cells those variant cells that might be used to immunize against the parenta

ISSN:0952-3499    

DOI:10.1002/jmr.300040406

出版商:John Wiley&Sons, Ltd.    

年代:1991

数据来源: WILEY

摘要:

AbstractA relatively recent method developed to determine the molecular weights of intact peptides and proteins, matrix‐assisted UV laser desorption time‐of‐flight mass spectrometry (LDTOF‐MS), has been evaluated as a new means to investigate the metal ion‐binding properties of model synthetic peptides. A contiguous sequence of 25 residues on the surface of the 74 kDA human plasma metal‐binding transport protein histidine‐rich glycoprotein (HRG) has been identified as a bioactive metal‐binding domain. The peptide, (GHHPH)5G, was synthesized and evaluated by LDTOF‐MS before and after the addition of Cu(II) in solution with 2,5‐dihydroxybenzoic acid as the matrix. In the absence of added Cu(II), the major protonated molecular ion (M + H)+was observed to have a mass equal to its calculated mass (2904.0 Da). In the presence of Cu(II), however, five additional peaks were observed at mass increments of approximately 63.9 Da. The maximum Cu(II)‐binding capacity observed for the 26‐residue peptide (5 g‐atoms/mol) suggest that up 1 Cu(II) may be bound per 5‐residue internal repeat unit (GHHPH) within this peptide: several other monovalent and divalent metal cations were not bound under identical conditions of analysis. The Cu(II)‐binding stoichiometry was verified by spectrophotometric titration and by frontal analyses of the immobilized peptide with a solution of Cu(II) ions. These results demonstrate the ability to verify directly the solution‐phase binding capacity of met

ISSN:0952-3499    

DOI:10.1002/jmr.300040407

出版商:John Wiley&Sons, Ltd.    

年代:1991

数据来源: WILEY

ISSN:0952-3499    

DOI:10.1002/jmr.300040401

出版商:John Wiley&Sons, Ltd.    

年代:1991

数据来源: WILEY