Computer analysis of protein structure.
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Alcohol Dehydrogenase↗
Biomedical subjects
P Argos
Publications and source records attributed to P Argos.
Sensitivity comparison of protein amino acid sequences.
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Amino Acid Sequence↗
Scrutineer: a computer program that flexibly seeks and describes motifs and profiles in protein sequence databases.
Scrutineer is an interactive, user-friendly program designed to search for motifs, patterns and profiles in the Swissprot, Protein Identification Resource (PIR) or SeqDb protein sequence databases. Basic capabilities include (i) searches for strings of amino acids with multiple choices at a given position; (ii) searches for strings including variable-length segments and delocalized constraints; (iii) searches over subsets of a database or particular regions within each sequence (e.g. N-terminal one-third); (iv) searches involving secondary structure predictions, physicochemical characteristics, and the like; and (v) searches using aligned sequences as targets with various optional weighting schemes. The various search criteria and hits can be combined and complex targets located. Once the data are loaded into virtual memory, all occurrences in PIR release 22.0 (3.7 x 10(6) amino acids) of a given short string of amino acids (e.g. a hexamer) are found in approximately 36 s. Scrutineer can also describe the entire database, user-specified hits, user-defined regions of sequence and all hits. The source code and accompanying manual are being freely distributed.
Algorithms↗
An attempt to unify the structure of polymerases.
With the great availability of sequences from RNA- and DNA-dependent RNA and DNA polymerases, it has become possible to delineate a few highly conserved regions for various polymerase types. In this work a DNA polymerase sequence from bacteriophage SPO2 was found to be homologous to the polymerase domain of the Klenow fragment of polymerase I from Escherichia coli, which is known to be closely related to those from Staphylococcus pneumoniae, Thermus aquaticus and bacteriophages T7 and T5. The alignment of the SPO2 polymerase with the other five sequences considerably narrowed the conserved motifs in these proteins. Three of the motifs matched reasonably all the conserved motifs of another DNA polymerase type, characterized by human polymerase alpha. It is also possible to find these three motifs in monomeric DNA-dependent RNA polymerases and two of them in DNA polymerase beta and DNA terminal transferases. These latter two motifs also matched two of the four motifs recently identified in 84 RNA-dependent polymerases. From the known tertiary architecture of the Klenow fragment of E. coli pol I, a spatial arrangement can be implied for these motifs. In addition, numerous biochemical experiments suggesting a role for the motifs in a common function (dNTP binding) also support these inferences. This speculative hypothesis, attempting to unify polymerase structure at least locally, if not globally, under the pol I fold, should provide a useful model to direct mutagenesis experiments to probe template and substrate specificity in polymerases.
Amino Acid Sequence↗
Determination of reliable regions in protein sequence alignments.
Judging the significance of alignments is still a major problem in sequence comparison. We present a method to delineate reliable regions within an alignment. This differs from standard approaches in that it does not attempt to attribute one significance value to the alignment as a whole, but assesses alignment quality locally. An algorithm is provided that predicts which residue pairs in an alignment are likely to be correctly matched. The predictions are evaluated by comparison with alignments taken from tertiary structural superpositions.
Algorithms↗
Effect of structural modifications on the assembly of a glycinin subunit.
A Gy4 glycinin cDNA was modified and used to produce structurally altered 11S storage protein subunits. We evaluated these modified subunits for their ability to assemble into oligomers. Alterations made in the acidic polypeptide changed the subunit solubility characteristics but did not eliminate assembly. Modifications in the basic polypeptide usually eliminated assembly of subunits into trimers. A region exhibiting high natural variability located at the COOH terminus of the acidic polypeptide that we have designated the hypervariable region was also studied. Extensive deletions and insertions were tolerated in the hypervariable region without perturbing subunit assembly. Some of the insertions significantly increased the methionine content in the Gy4 glycinin subunit. Together, our results indicated that the structure of the basic polypeptide was more critical for assembly of trimers than that of the acidic polypeptide, an observation that implies that the basic polypeptides direct trimer formation. The assembly assays described here will be useful in efforts to improve seed quality. Using them, the effects of modifications to the storage protein subunits can be rapidly evaluated before introducing the mutated genes into plants.
Base Sequence↗
The primary structure of human secretogranin II, a widespread tyrosine-sulfated secretory granule protein that exhibits low pH- and calcium-induced aggregation.
Secretogranin II (previously also called chromogranin C) is a tyrosine-sulfated secretory protein found in secretory granules in a wide variety of endocrine cells and neurons. Here, we have determined the primary structure of human secretogranin II from a full length cDNA clone and have investigated its properties, predicted from the sequence, by studying the behavior of purified secretogranin II under conditions characteristic of the milieu of secretory granules. Analysis of a 2.35-kilobase cDNA clone isolated from a human pituitary library and identified as secretogranin II by various criteria showed that human presecretogranin II is a 617-residue polypeptide containing an NH2-terminal located signal peptide. Secretogranin II lacks the disulfide-bonded loop structure near the NH2 terminus which is conserved in chromogranin A and chromogranin B (secretogranin I), two other widespread constituents of neuroendocrine secretory granules, but like the latter two proteins contains (i) an -E-N/S-L-X-A/D-X-D/E-X-E-L- motif and (ii) multiple potential dibasic cleavage sites for the generation of smaller, perhaps biologically active peptides. Another structural feature that secretogranin II shares with chromogranin A and chromogranin B (secretogranin I) is the abundance of acidic residues all along the polypeptide chain whose negative charge must somehow be neutralized to allow condensation and packaging of the protein into secretory granules. Experiments with purified secretogranin II showed that in the presence of 10 mM calcium at pH 5.2, conditions characteristic of the milieu of neuroendocrine secretory granules, this protein formed aggregates. Immunoglobulin G, a secretory protein that in vivo is not packaged into secretory granules, did not form aggregates under these in vitro conditions and was excluded from the secretogranin II aggregates. Very little aggregation of secretogranin II was observed in the absence of calcium at pH 5.2 or in the presence of calcium at neutral pH. In vivo, ammonium chloride, which is known to neutralize the pH of acidic intracellular compartments, inhibited the packaging of newly synthesized secretogranin II into secretory granules. Our results suggest that the low pH- and calcium-induced aggregation of secretogranin II may be important for the organization of the secretory granule matrix and raise the possibility that aggregation of secretogranin II may be involved in its sorting to secretory granules.
Amino Acid Sequence↗
Engineering protein thermal stability. Sequence statistics point to residue substitutions in alpha-helices.
Amino acid sequences have been compared for thermophilic and mesophilic molecules from six different protein families, which include lactate and glyceraldehyde-3-phosphate dehydrogenases, triose phosphate isomerases, superoxide dismutases, thermolysins and subtilisins. Since a three-dimensional structure was known for at least one of the sequences in each family, analysis of preferred residue substitutions, presumably to achieve thermal stability, could be examined from a structural context. The overall results, which are generally consistent across all the families, suggested decreased flexibility and increased hydrophobicity in alpha-helical regions as the main stabilizing principles. The most favoured residual exchanges, hopefully useful in engineering stability into proteins, are discussed.
Amino Acid Sequence↗
A possible homology between immunodeficiency virus p24 core protein and picornaviral VP2 coat protein: prediction of HIV p24 antigenic sites.
With the use of a sensitive sequence comparison algorithm, a homology has been suggested between the primary structures of simian immunodeficiency virus (SIV) p24 core protein and foot-and-mouth disease virus (FMD) VP2 coat protein. Since the FMD sequence is homologous to picornaviral VP2 sequences with known three-dimensional architecture and since the SIV p24 sequence can be convincingly aligned with that from human immunodeficiency virus (HIV), it was possible to predict an eight-stranded beta-barrel fold for the HIV core protein. From analogy with the known environments of the picornaviral coats, p24 sequence spans could be predicted as likely candidates for antibody attachment. These suggestions may be important for development of an AIDS vaccine.
Amino Acid Sequence↗
A new interactive protein sequence alignment program and comparison of its results with widely used algorithms.
A computer program that allows interactive sequence comparison is described. It graphically displays a search matrix using residue physiochemical characteristics and multilength segmental comparisons. The user selects through a mousing device and screen pointer the sequence spans to be matched. The results of this method are compared with those of ALIGN and BESTFIT.
Algorithms↗
A fast and sensitive multiple sequence alignment algorithm.
A two-step multiple alignment strategy is presented that allows rapid alignment of a set of homologous sequences and comparison of pre-aligned groups of sequences. Examples are given demonstrating the improvement in the quality of alignments when comparing entire groups instead of single sequences. The modular design of computer programs based on this algorithm allows for storage of aligned sequences and successive alignment of any number of sequences.
Algorithms↗
A sequence motif in many polymerases.
A 15-residue sequence motif has been found in many polymerases from various species and involving DNA and RNA dependence and product. The motif is characterized by a Tyr-Gly-Asp-(Thr)-Asp core flanked by hydrophobic spans five residues in length. An mRNA maturase segment is also suggested to display the motif pattern. The aspartates may be important in polymerase function by acting directly in catalysis and/or by binding magnesium.
Amino Acid Sequence↗
Unexpected divergence and molecular coevolution in yeast plasmids.
Four closely related species of yeast possess multicopy nuclear plasmids whose shared molecular architecture demonstrates a common ancestor, despite their lack of discernible DNA sequence homology. Each plasmid encodes three proteins which have equivalent essential functions in plasmid maintenance. These three groups of proteins show markedly different degrees of conservation, so that although we have successfully aligned sequences for two groups, members of the third group have diverged to such an extent that they cannot be aligned. All the proteins are sufficiently different that they function only in conjunction with their encoding plasmid. These proteins have therefore conserved their functional interactions with the relevant DNA sequences of their particular plasmids, despite lack of amino acid sequence conservation. The maintenance of function in the face of DNA sequence divergence is analogous to the coevolution of ribosomal DNA promoters and RNA polymerase I, and suggests that molecular drive may be an important force in the evolution of these plasmids. This view is reinforced by the inconsistent phylogenetic relationships determined from the two alignment sets, and by the contradiction that the two plasmids known to be the closest related taxonomically and by their host interchangeability are suggested to be the most distant by their sequences.
Amino Acid Sequence↗
Sugar permeases of the bacterial phosphoenolpyruvate-dependent phosphotransferase system: sequence comparisons.
The amino acyl sequences of eight permeases (enzymes II and enzyme II-III pairs) of the bacterial phosphoenolpyruvate:sugar phosphotransferase system (PTS) have been analyzed. All systems show similar sizes, and six of these systems exhibit the same molecular weight +/- 2%. Several exhibit sequence homology. Characteristic NH2-terminal and COOH-terminal sequences were found. The NH2-terminal leader sequences are believed to function in targeting of the permeases to the membrane, whereas the characteristic COOH-terminal sequences are postulated to mediate interaction with the energy-coupling protein phospho HPr. One of the systems, the one specific for mannose, exhibits distinctive characteristics. A pair of probable phosphorylation sites was detected in each of the five most similar systems, those specific for beta-glucosides, sucrose, glucose, N-acetylglucosamine, and mannitol. One of the two equivalent phosphorylation sites (proposed phosphorylation site 1) was located approximately 80 residues from the COOH terminus of each system. The other site (proposed phosphorylation site 2) was located approximately 440 residues from the COOH termini of the glucose and N-acetylglucosamine systems, approximately 320 residues from the COOH termini of the beta-glucoside and sucrose systems, and 381 residues from the COOH terminus of the mannitol system. Intragenic rearrangement during evolutionary history may account for the different positions of phosphorylation sites 2 in the different PTS permeases. More extensive intragenic rearrangements may have given rise to entirely different positions of phosphorylation in the glucitol, mannose, and lactose systems. A single, internal amphipathic alpha-helix with characteristic features was found in each of seven of the eight enzymes II. The lactose-specific enzyme III of Staphylococcus aureus was unique in possessing a COOH-terminal amphipathic alpha-helix rich in basic amino acyl residues. Possible functions for these amphipathic segments are discussed.
Amino Acid Sequence↗
A model for the hepatitis B virus core protein: prediction of antigenic sites and relationship to RNA virus capsid proteins.
The sequences of the core proteins from several serotypes of human hepatitis B virus and related mammalian and avian hepadnaviruses are aligned with the vp3 capsid protein of mengo virus, a picornavirus. The homology indicates an eight-stranded antiparallel beta-barrel fold for the hepatitis protein, as observed in the tertiary structure of the picornavirus protein. The locations of known antigenic sites and other modifications are consistent with this structure for the core protein. The predicted folding suggests additional exposed antigenic sites and supports an evolutionary relationship between this family of enveloped DNA viruses and enveloped and non-enveloped RNA viruses.
Capsid↗
Oligopeptide biases in protein sequences and their use in predicting protein coding regions in nucleotide sequences.
We have examined oligopeptides with lengths ranging from 2 to 11 residues in protein sequences that show no obvious evolutionary relationship. All sequences in the Protein Identification Resource database were carefully classified by sensitive homology searches into superfamilies to obtain unbiased oligopeptide counts. The results, contrary to previous studies, show clear prejudices in protein sequences. The oligopeptide preferences were used to help decide the significance of sequence homologies and to improve the more general methods for detecting protein coding regions within nucleotide sequences.
Amino Acid Sequence↗
An investigation of protein subunit and domain interfaces.
Protein structures were collected from the Brookhaven Database of tertiary architectures that displayed oligomeric association (24 molecules) or whose polypeptide folding revealed domains (34 proteins). The subunit and domain interfaces for these proteins were respectively examined from the following aspects: percentage water-accessible surface area buried by the respective associations, surface compositions and physical characteristics of the residues involved in the subunit and domain contacts, secondary structural state of the interface amino acids, preferred polar and non-polar interactions, spatial distribution of polar and non-polar residues on the interface surface, same residue interactions in the oligomeric contacts, and overall cross-section and shape of the contact surfaces. A general, consistent picture emerged for both the domain and subunit interfaces.
Amino Acids↗
A model for the tertiary structure of the 28 residue DNA-binding motif ('zinc finger') common to many eukaryotic transcriptional regulatory proteins.
Many eukaryotic transcriptional activator proteins, including the Xenopus 5S RNA gene activator protein TFIIIA and the HeLa cell protein Sp1, have an approximately 30 amino acid repeating motif which binds to short, specific DNA sequences. Over 150 of these sequences are now known. Based on the observed distribution of amino acid residues, a series of constraints and predictions can be proposed for the structure of the motif. A compatible three-dimensional structural model has been developed by a combination of interactive model building and refinement by molecular dynamics. The model structure consists of a two-stranded beta-hairpin stabilizing a C-terminal alpha-helix by both zinc ligands and hydrophobic interactions. Four of the residue positions on the helix N-terminus and exposed face are predicted to provide base specific ligands. Further implications of the model for DNA binding are discussed.
Amino Acid Sequence↗