摘要:

AbstractAn attempt has been made to build a model of human rhinovirus 2 (HRV2) based on the known human rhinovirus 14 (HRV14) structure. HRV2 was selected because its amino acid sequence is known and because it belongs to the minor rhinovirus receptor class as compared to HRV14, which belongs to the major class. Initial alignment of HRV2 with HRV14 based on the primary sequence and the knowlege of the three‐dimensional structure of HRV14 showed that most probable position of the majority of insertions and deletions occurred in the vicinity of the neutralizing immunogenic sites (NIm). Out of a total of 855 amino acids present in one copy of each of the capsid proteins VP1 through VP4 of HRV14, 411 are different between the two viruses. There are also 6 amino acids residues inserted and 14 residues deleted in HRV2 relative to HRV14. Examination of amino acid interactions showed several cases of conservation of function, e.g., salt bridges or the filling of restricted space. The largest variation amongst the residues lining the canyon, the putative receptor binding site, was in the carboxy‐terminal residues of

ISSN:0887-3585    

DOI:10.1002/prot.340020402

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

摘要:

AbstractEcoRI endonuclease mutants were isolated in a methylase‐deficient background following in vitro hydroxylamine mutagenesis of plasmid pKG2 (Kuhn et al.:Gene44:253–263, 1986). Mutants which survived high‐level endonuclease expression (IPTG induction) were termed null mutants. Sixtytwo of 121 null mutants tested by Western blot contained normal levels of endonuclease cross‐reacting protein. The complete endonuclease gene was scquenced for 27 null mutants. This group was found to consist of 20 signle base‐change missense mutations, 6 double mutations, and 1 triple mutation. Ten of the 20 signle mutations were clustered between residues 139 and 144. When examined with respect to the structure of theEcoRI‐DNA complex (McClarin et al.:Science234:1526–1541, 1986), these alterations werre found to fall predominantly into two classes: substitutions at the protein‐DNA interface or substitutions at the protein‐protein (dimer) interface. Protein from several of the mutants was purified and sized by using HPLC. Wild‐typeEcoRI endonuclease and protein from three of the DNA interface mutations (A1a139→Thr, Gly140→Ser, Arg203→Gln) appeared to be dimeric, while protein from subunit interface mutations (Glu144→Lys, Glu152→Lys, Gly

ISSN:0887-3585    

DOI:10.1002/prot.340020403

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

摘要:

AbstractTwo observations suggest that DNA, upon binding toE. colicatabolite gene activator protein (CAP), is sharply bent by a total angle of at least 100–150 degrees: (1) The electrostatic protential field of CAP shows regions of positive potential that from a ramp on 3 sides of the protein. (2) The DNA binding site size as determined by DNA ethylation interference with binding, (Majors: “Control of theE. coliLac Operon at the Molecular Level.” Ph.D. Thesis, Harvard University, Cambridge, 1977) and by relative affinities of DNA fragments of various lengths (Liu‐Johnson et al.:Cell47:995–1005, 1986) requires severe bending of the DNA to maintain its favorable electrostatic contact with th

ISSN:0887-3585    

DOI:10.1002/prot.340020404

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

摘要:

AbstractUniquely among class A β‐lactamases, the RTEM‐1 and RTEM‐2 enzymes contain a single disulfide bond between Cys 77 and Cys 123. To study the possible role of this naturally occurring disulfide in stabilizing RTEM‐1 β‐lactamase and its mutants at residue 71, this bond was removed by introducing a Cys 77 → Ser mutation. Both the wild‐type enzyme and the single mutant Cys 77 → Ser confer the same high levels of resistance to ampicillin in vivo toEcherichia coli; at 30°C the specific activity of purified Cys 77 → Ser mutant is also the same as that of the wild‐type enzyme. Also, neither wild‐type enzyme nor the Cys 77 → Ser mutant is inactivated by brief exposure to p‐hydroxymercuri‐benzoate. However, above 40°C the mutant enzyme is less stable than wild‐type enzyme. After introduction of the Cys 77 → Ser mutation, none of the double mutants (containing the second mutations at residue 71) confer resistance to ampicillin in vivo at 37°C; proteins with Ala, Val, Leu, Ile, Met, Pro, His, Cys, and Ser at residue 71 confer low levels of resistance to ampicillin in vivo at 30°C. The use of electrophoretic blots stained with antibodies against β‐lactamase to analyze the relative quantities of mutant proteins in whole‐cell extracts ofE. colisuggests that all 19 of the doubly mutant enzymes are proteolyzed much more readily than their singly mutant analogues (at Thr 71) that contain a disulfide bond. Thus, the disulfide bond of the RTEM‐1 β‐lactamase seems able to reduce the destabilizing effect of mutations at Thr 71. These results also emphasize the unique and essential role that Thr 71 performs in the stable folding of RTEM‐1 β‐lac

ISSN:0887-3585    

DOI:10.1002/prot.340020405

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

摘要:

AbstractPeptides corresponding to portions of loop 2 of snake venom curare‐mimetic neurotoxins and to a structually similar region of rabies virus glycoprotein were synthesized. Interaction of these peptides with purifiedTorpedoelectric organ acetylcholine receptor was tested by measuring their ability to block the binding of125I‐labeled α‐bungarotoxin to the receptor. In addition, inhibition of α‐bungarotoxin binding to a 32‐residue synthetic peptide corresponding to positions 173–204 of the α‐subunit was determined. Neurotoxin and glycoprotein peptides corresponding to toxin loop 2 inhibited labeled toxin binding to the receptor with IC50values comparable to those of nicotine and the competitive antagonist d‐tubocurarine and to the α‐subunit peptides with apparent affinities between those of d‐tubocurarine and α‐cobratoxin. Substitution of neurotoxin residue Arg37, the proposed counterpart of the quatenary ammonium of acetylcholine, with a negatively charged Glu residue reduced the apparent affinity about 10‐fold. Peptides containg the neutotoxin invarint residue Trp29 had 10‐ to 100‐fold higher affinites than peptides lacking this residue. These results demonstrate that relatively short synthetic peptides retain some of binding ability of the native protein from which they are derived, indicating that such peptides are useful in the study of proetin interactions. The ability of the peptides to compete α‐bungarotoxin binding to the receptor with apparent affinites comparable to those of other cholinergic ligands indicates that loop 2 of the neurotoxins and the strucually similar segment of the rabies virus glycoprotein act as recognition sites for the acetylcholine receptor. Invarient toxin residues Arg37 and Trp29 and their viral homologs play important, although not essential, roles in binding, possibly by interaction with complementary anionic and hydrophobic subsites on the acetylcholine receptor. The α‐subunit peptide most likely contains all of the determinants for binding of the toxin and glycoprotein peptides present on the α‐subunit, because these peptides bind to the 32‐residue α‐subunit peptides with the sam

ISSN:0887-3585    

DOI:10.1002/prot.340020406

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

摘要:

AbstractThe normal mode analysis of conformational fluctuation is carried out for a small globular protein, bovine pancreatic trypsin inhabitor. Results are analyzed mainly to reveal the mechanical construction of the protein molecule. We take dihedral angles, including peptide ω angles, as independent variables for the normal mode analysis. There are 306 such angles in this molecule. Motions in modes with frequencies lower than 120 cm−1are showned to involve atoms in the whole protein molecule, and spatial change of displacement vectors is continues, i.e., those of atoms near in space are similar. To quantitate the observation of the continuity, a correlation function of direction vectors of atomic displacement is calculated. From this function we define a Quantity that is interpreted as the wave length of an equivalent elastic plane wave. From this Quantity we deduce eefective young's modulus for each mode. For the mode with the lowest frequency 4.4 cm−1, it terned out to be 0.8 × 109dyn cm−2, the value two orders magnitude softer than, for instance, α‐helices. Prompted by this observation, the four lowest frequency modes and also the harmonic motions in the thermal equilibrium are analyzed further mainly to detect relatively rigid structural elements in the molecule. From this analysis emerges a mechanical picture of the protein molecule that is made up of relatively rigid elements held together by very

ISSN:0887-3585    

DOI:10.1002/prot.340020407

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

摘要:

AbstractEffective van der Waals radii were celebrated in such a way that molecular models built from standard bond lengths and bond angels reproduced the amino acid conformations observed by crystallography in proteins and peptides. The celebrations were based on the comparision of the Ramachandbran plots prepared from high‐resolution X‐ray data of protins and peptides with the allowed ϕ,ψ torsional angel space for the depeptide molecular models. The celebrated radii are useful as criteria with which to filter energetically improbable conformations in molecular modeling studies of proteins and pep

ISSN:0887-3585    

DOI:10.1002/prot.340020408

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

摘要:

AbstractA new method for the analysis of NMR data in terms of the solution structure of proteins has been developed. The method consists of two steps: first systematic search of the conformational space to define the region allowed by the initial set of experimental constraints, and second, the narrowing of this region by the introduction of aaditional constraint and optional refinement procedures. The search of the conformational space is guided by heuristics to make it computationally feasible. The method is therfore called the heuristic refinement method and is coded in an expert system called PROTEAN. The paper describe the validation of the first step of the method using an artificial NMR data set generated from the known crystal structure of sperm whale carbon monoxymyoglodin. It is shown that the initial search procedure yields a low‐resolution structure of the myoglobin molecule, accurately reproducing its main topological features, and that the precision of the structure depend on the quality of the intial data se

ISSN:0887-3585    

DOI:10.1002/prot.340020409

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY

ISSN:0887-3585    

DOI:10.1002/prot.340020401

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1987

数据来源: WILEY