摘要:

AbstractAlzheimer's disease is characterized by the deposition of amyloid β‐protein as plaques and tangles in the brains of its victims. The amyloid precursor can be expressed with or without the inclusion of a protease inhibitor domain, the potential role of which in amyloidogenesis has prompted the generation of a model of its three‐dimensional structure based on the known structure of a related inhibitor. The model structure predicts that the mutated residues are almost entirely on the surface of the inhibitor domain, while conserved residues constitute the hydrophobic core. In addition, several pairs of structurally complementary, or concerted, mutations are seen. These structural features provide strong evidence for the validity of the modeled structure, and it is suggested that the presence of complementary mutations may be used as a criterion for evaluating protein structures built by homology, in addition to the (spatial) location of the mutations. The terminal residues delimiting the domain are among those furthest from the protease binding site and are in close proximity to one another, thus suggesting the ability of the domain to function as a structural cassette within the context of a larger protein. The electrostatic potentials of the inhibitor and of the related bovine pancreatic trypsin inhibitor reveal how two inhibitors with very different net charges can bind with approximately the same binding constant to trypsin and suggest a mutation of trypsin that might selectively enhance the binding of the amyloid inhibitor domain. The model provides a structural basis for understanding the functional roles of residues in the domain and for designing simpler molecules to test as pharmacologic agents for intervention in Alzheimer's dis

ISSN:0887-3585    

DOI:10.1002/prot.340090102

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

年代:1991

数据来源: WILEY

摘要:

AbstractA series of 37‐residue analogues of the pseudo‐EF hand in bovine calbindin D9Khas been synthesized by the solid phase method. In the presence of calcium an α‐helical induction of up to 44% was observed for the peptide with the native sequence with aKdfor calcium binding of 0.35 mM. A number of amino acid substitutions have been carried out to study the packing of the two α‐helices based on the crystal structure of the entire protein. Three strategies were employed: (1) replacement of the Leu residues, which in the crystal structure do not contribute to the hydrophobic interaction between the two helices, by Gln or Ala in order to control the orientation of the helix packing, (2) stabilization of the individual helix by introducing a Glu−…︁Lys+salt bridge or by changing the N‐terminal charge to compensate for the helix dipole moment, and (3) introduction of a disulfide bond between the two helices to help the packing of the helices.The mutants with the substitution of (Leu‐30, Leu‐32) to (Gln‐30, Gln‐32), (Gln‐30, Ala‐32), and (Ala‐30, Ala‐32) designed based on the strategy 1 do not show any affinity for calcium and have low α‐helicity. The Leu‐30 to Lys‐30 mutant designed to form a salt bridge between the side chains of Glu‐26 and Lys‐30 has an apparentKdfor calcium of 6.8 mM.Kdof the N‐terminal acetylated and succinylated mutants are 0.41 and 0.45 mM, respectively, and no increase in the α‐helix content relative to that of the natural sequence peptide is observed. The disulfide containing mutants, namely Tyr‐13, Leu‐31 to Cys‐31 and Tyr‐13, Leu‐31 to Cys‐13, hCys‐31, show apparentKdvalues of 0.93 and 2.1 mM, respectively. The former mutant shows the highest α‐helix content among the peptides studied in the presence and absence of calcium. While it is difficult to construct an isolated and rigid helix–loop–helix motif with peptides of this size, in

ISSN:0887-3585    

DOI:10.1002/prot.340090103

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

年代:1991

数据来源: WILEY

摘要:

AbstractAn explanation is suggested for why a marginally stable β‐structure folds extremely slowly; it is predicted that even a small increase in stability drastically accelerates β‐folding. According to the theory, this folding is a first‐order phase transition, and the rate‐limiting step is nucleation. The rate‐determining “nucleus” (transition state) is the smallest β‐sheet that is sufficiently large to provide an overall free energy reduction during subsequent folding. If the stability of the β‐structure is low, the nucleus is large and possesses a high free energy due to having a large perimeter. When the net stability of the final β‐structure increases (due to either an increase of the β‐sheet stability or a decrease in stability of the competing structures, e.g., α‐helices), the size and energy of a nucleus decrease and the rate of folding increases exponentially. This must result in a fast folding of polypeptides enriched by β‐forming residues (e.g., protein chains). The theory is developed for intramolecular β‐structure, but it can also explain the overall features of intermolecular β‐folding; it is applicable both to antiparallel and parallel β‐sheets. The difference in folding of β

ISSN:0887-3585    

DOI:10.1002/prot.340090104

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

年代:1991

数据来源: WILEY

摘要:

AbstractSite‐directed mutagenesis of key amino acids of adenylate kinase has been used to suggest a new model for the location of the AMP and ATP binding sites. Phe‐86 and Tyr‐133, which are in close contact with the inhibitor Ap5A according to previous crystallographic results, have been independently changed to tryptophan and other amino acids. The Phe‐86→Trp mutant had a 3‐ to 6‐fold change in theKmfor ATP and a 44‐fold increase in theKmfor AMP with a simultaneous loss of AMP substrate inhibition. Thus Phe‐86 is probably in close contact with bound AMP. The Tyr‐133→Trp mutant showed no large effects on enzyme kinetics and suggests that the previous assignment of Ap5A occupying natural adenosine binding sites is probably incorrect. A temperature‐sensitive Leu‐107→Gln mutant showed a 6‐fold decrease in theKmfor ATP and no effect on AMP binding, suggesting that this amino acid is near the ATP binding site.Changes in the fluorescence of single tryptophan‐containing mutant enzymes provided specific information about AMP and ATP binding. The fluorescence results are consistent with the kinetic studies, and also suggest that AMP substrate inhibition is caused by the formation of an abortive complex that

ISSN:0887-3585    

DOI:10.1002/prot.340090105

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

年代:1991

数据来源: WILEY

摘要:

AbstractA survey of 40 multisubunit proteins and 2 protein–protein complexes was performed to assay quantitatively the distribution of hydropathy among the exterior surface, interior, contact surface, and noncontact exterior surface of the isolated subunits. We suggest a useful way to present this distribution by using a “hydropathy level diagram.” Additionally, we have devised a function called “hydropathy complementarity” to quantitate the degree to which interacting surfaces have matching hydropathy distributions. Our survey revealed the following patters: (1) The difference in hydropathy between the interior and exterior of subunits is a fairly invariant quantity. (2) On average, the hydropathy of the contact surface is higher than that of the exterior surface, but is not greater than that of the protein as a whole. There was variation, however, among the proteins. In some instances, the contact surface was more hydrophilic than the noncontact exterior, and in a few cases the contact surface was as hydrophobic as the protein interior. (3) The average interface manifests significant hydropathy complementarity, signifying that proteins interact by placing hydrophobic centers of one surface against hydrophobic centers of the other surface, and by similarly matching hydrophilic centers. As a measure of recognition and specificity, hydropathy complementarity could be a useful tool for predicting correct docking of interacting proteins. We suggest that high hydropathy complementarity is associated with static inflexible interactions. (4) We have found that some subunits that bind predominantly through hydrophilic forces, such as hydrogen bonds, ionic pairs, and water and metal bridges, are involved in dynamic quaternary organization and

ISSN:0887-3585    

DOI:10.1002/prot.340090106

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

年代:1991

数据来源: WILEY

摘要:

AbstractThe database of known protein three‐dimensional structures can be significantly increased by the use of sequence homology, based on the following observations. (1) The database of known sequences, currently at more than 12,000 proteins, is two orders of magnitude larger than the database of known structures. (2) The currently most powerful method of predicting protein structures is model building by homology. (3) Structural homology can be inferred from the level of sequence similarity. (4) The threshold of sequence similarity sufficient for structural homology depends strongly on the length of the alignment. Here, we first quantify the relation between sequence similarity, structure similarity, and alignment length by an exhaustive survey of alignments between proteins of known structure and report a homology threshold curve as a function of alignment length. We then produce a database of homology‐derived secondary structure of proteins (HSSP) by aligning to each protein of known structure all sequences deemed homologous on the basis of the threshold curve. For each known protein structure, the derived database contains the aligned sequences, secondary structure, sequence variability, and sequence profile. Tertiary structures of the aligned sequences are implied, but not modeled explicity. The database effectively increases the number of known protein structures by a factor of five to more than 1800. The results may be useful in assessing the structural significance of matches in sequence database searches, in deriving preferences and patterns for structure prediction, in elucidating the structural role of conserved residues, and in modeling three‐dimensional detail by hom

ISSN:0887-3585    

DOI:10.1002/prot.340090107

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

年代:1991

数据来源: WILEY

摘要:

AbstractPatterns in amino acid properties (polar, hydrophobic, etc.) that characterize secondary structure motifs are derived from a database containing 75 protein structures, with the aim of circumventing the limitations due to data base size so as to increase structure prediction score. Many such sequence–structure associations with high intrinsic predictive power are found, which turn out to be correct 78% of the time when applied individually to proteins outside the learning set. Based on these associations, a prediction method is developed, which reaches the socre of 62% on the 3 states α‐helix, β‐strand, and loop, without using additional constraints. Though this score is quite good compared to that of other available prediction methods, it is much lower than could be expected from the high intrinsic predictive power of the associations used. The reasons underlying this surprising result, which indicate that prediction score and intrinsic predictive power are only weakly coupled, are discussed. It is also shown that the size of the present database still seriously limits prediction scores, even when property patterns are used, and that higher scores are expected in large databases. Clues are provided on the relative influence of neglecting spatial interactions on prediction efficiency, suggesting that, in sufficiently large databases, predicted secondary structures would correspond to those formed early in the folding process. This hypothesis is tested by confronting present predictions with available experimental data on early protein folding intermediates and on small peptides that adopt a relatively stable conformation in water. Although admittedly there are still too few such data, results suggest that the hypothesis might be well

ISSN:0887-3585    

DOI:10.1002/prot.340090108

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

年代:1991

数据来源: WILEY

ISSN:0887-3585    

DOI:10.1002/prot.340090109

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

年代:1991

数据来源: WILEY

ISSN:0887-3585    

DOI:10.1002/prot.340090101

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

年代:1991

数据来源: WILEY