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S Pascarella

Publications and source records attributed to S Pascarella.

32 records · Page 2Linked to original sources
PubMed1994

Conservation of amphipathic conformations in multiple protein structural alignments.

Protein amphipathic conformations, mainly alpha-helices and beta-strands, are believed to play an important role in protein folding, stability and function. The most popular method for characterizing such structures is the hydrophobic moment. We have analyzed the distribution of hydrophobic moment characteristics (peak magnitude, amphipathic indices and characteristic frequency) in a data bank containing several families of distant sequences multiply aligned by structural superposition. Sequence fragments were classified according to alpha-helix, beta-strand, non-alpha and non-beta conformations. This data bank provided an enhanced sample space compared with those previously reported in the literature. Precautions were taken to reduce over-representation of homologous sequences. Approximately 50% of all individual alpha-helices showed a hydrophobic moment peak in the expected position of the periodicity spectrum while only 38% of individual beta-strands fell in the expected range. False positives account for a surprisingly large 14 and 36% of the non-alpha and non-beta samples respectively. Conservation of hydrophobic moment characteristics and mainly the hydrophobic peak position in the expected periodicity range was examined in the multiple alignments of the distant sequences. Helices tend to conserve more frequently their hydrophobic moment than any other conformation and yet only 13% of all helical segments display such conservation in three-quarters or more of the familial sequences; the similar observation for beta-strands was even lower at 9%. Nonetheless, strongly hydrophobic positions within the structural segments were more conserved than expected.

Amino Acid Sequence↗
PubMed1993

Similarity between serine hydroxymethyltransferase and other pyridoxal phosphate-dependent enzymes.

A structural homology of the pyridoxal-5'-phosphate (PLP)-dependent enzyme serine hydroxymethyltransferase (SHMT) with aspartate aminotransferase (AAT) is proposed. Although the two sequences are very dissimilar, a reasonable alignment was obtained using the profile analysis method. Sequences of AAT and dialkylglycine decarboxylase (DGD), for which crystal structure data are available, have been aligned on the basis of their structure superposition. A profile was then calculated and SHMT sequence aligned to it. Three of the four residues conserved in all aminotransferases (including the PLP-binding lysine) are matched. A profile search with DGD-AAT-SHMT profile is more selective and sensitive than individual sequence profiles for PLP-dependent enzyme detection. Potential homologies with the eryC1 gene product involved in erythromycin biosynthesis and with amino acid decarboxylases were observed. Homology with AAT will be used as a guideline for planning site-directed mutagenesis experiments on SHMT.

Amino Acid Sequence↗
PubMed1993

Quantification of secondary structure prediction improvement using multiple alignments.

The use of multiple sequence alignments for secondary structure predictions is analysed. Seven different protein families, containing only sequences of known structure, were considered to provide a range of alignment and prediction conditions. Using alignments obtained by spatial superposition of main chain atoms in known tertiary protein structures allowed a mean of 8% in secondary structure prediction accuracy, when compared to those obtained from the individual sequences. Substitution of these alignments by those determined directly from an automated sequence alignment algorithm showed variations in the prediction accuracy which correlated with the quality of the multiple alignments and distance of the primary sequence. Secondary structure predictions can be reliably improved using alignments from an automatic alignment procedure with a mean increase of 6.8%, giving an overall prediction accuracy of 68.5%, if there is a minimum of 25% sequence identity between all sequences in a family.

Amino Acid Sequence↗
PubMed1992

Analysis of insertions/deletions in protein structures.

An analysis of insertions and deletions (indels) occurring in a databank of multiple sequence alignments based on protein tertiary structure is reported. Indels prefer to be short (1 to 5 residues). The average intervening sequence length between them versus the percentage of residue identity in pairwise alignments shows an exponential behaviour, suggesting a stochastic process such that nearly every loop in an ancestral structure is a possible target for indels during evolution. The results also suggest a limit to the average size of indels accommodated by protein structures. The preferred indel conformations are reverse turn and coil as are the preferred conformations at the indel edges (N- and C-terminal sides). Interruptions in helices and strands were observed as very rare events.

Amino Acid Sequence↗
PubMed1992

Serine hydroxymethyltransferase: origin of substrate specificity.

All forms of serine hydroxymethyltransferase, for which a primary structure is known, have five threonine residues near the active-site lysyl residue (K229) that forms the internal aldimine with pyridoxal phosphate. For Escherichia coli serine hydroxymethyltransferase each of these threonine residues has been changed to an alanine residue. The resulting five mutant enzymes were purified and characterized with respect to kinetic and spectral properties. The mutant enzymes T224A and T227A showed no significant changes in kinetic and spectral properties compared to the wild-type enzyme. The T225A and T230A enzymes exhibited differences in Km and kcat values but exhibited the same spectral properties as the wild-type enzyme. The four threonine residues at positions 224, 225, 227, and 230 do not play a critical role in the mechanism of the enzyme. The T226A enzyme had nearly normal affinity for substrates and coenzymes but had only 3% of the catalytic activity of the wild-type enzyme. The spectrum of the T226A enzyme in the presence of amino acid substrates showed a large absorption maximum at 343 nm with only a small absorption band at 425 nm, unlike the wild-type enzyme whose enzyme-substrate complexes absorb at 425 nm. Rapid reaction studies showed that when amino acid substrates and substrate analogues were added to the T226A enzyme, the internal aldimine absorbing at 422 nm was rapidly converted to a complex absorbing at 343 nm in a second-order process. This was followed by a very slow first-order formation of a complex absorbing at 425 nm.(ABSTRACT TRUNCATED AT 250 WORDS)

Alanine↗
PubMed1992

A data bank merging related protein structures and sequences.

A data collection which merges protein structural and sequence information is described. Structural superpositions amongst proteins with similar main-chain fold were performed or collected from the literature. Sequences taken from the protein primary structure databases were associated with the multiple structural alignments providing they were at least 50% homologous in residue identity to one of the structural sequences and at least 50% of the structural sequence residues were alignable. Such restrictions allow reasonable confidence that the primary sequences share the conformation of the tertiary structural templates, except in the less conserved loop regions. Multiple structural superpositions were collected for 38 familial groups containing a total of 209 tertiary structures; 45 structures had no superposable mates and were used individually. Other information is also provided as main-chain and side-chain conformational angles, secondary structural assignments and the like. Wedding the primary and tertiary structural data resulted in an 8-fold increase of data bank sequence entries over those associated with the known three-dimensional architectures alone.

Amino Acid Sequence↗
PubMed1989

The primary structure of rabbit liver mitochondrial serine hydroxymethyltransferase.

The complete amino acid sequence of mitochondrial serine hydroxymethyltransferase from rabbit liver was determined. The sequence was obtained from analysis of peptides isolated from chymotryptic, cyanogen bromide, and limited acid cleavages of the protein. The enzyme consists of four identical subunits, each of 475 residues, i.e. 8 residues shorter than the subunit of the corresponding cytosolic isoenzyme. The sequences of the two rabbit proteins are easily aligned, provided a gap of 5 residues near the amino terminus and a gap of 3 residues near the carboxyl terminus are included in the mitochondrial sequence. The overall degree of identity between the two isoenzymes is 61.9%, whereas the structural identity of each eukaryotic isoenzyme with the corresponding Escherichia coli enzyme is about 40%. The rabbit isoenzymes are about 70 residues longer than the E. coli enzyme, with one-half of these residues accounted for by insertions in both isoenzymes near their carboxyl terminus. Predictions of secondary structure and calculations of hydropathy profiles are also presented, suggesting an even more extensive degree of identity in the three-dimensional folding of the three proteins, in accord with the known similarity of their catalytic properties. Evidence was obtained for the existence of additional molecular forms of the mitochondrial protein, differing in the absence of some amino acid residues at the amino terminus of the polypeptide chain.

Amino Acid Sequence↗
PubMed1987

The primary structure of rabbit liver cytosolic serine hydroxymethyltransferase.

The complete amino acid sequence of cytosolic serine hydroxymethyltransferase from rabbit liver was determined. The sequence was determined from analysis of peptides isolated from tryptic and cyanogen bromide cleavages of the enzyme. Special procedures were used to isolate and sequence the C-terminal and blocked N-terminal peptides. Each of the four identical subunits of the enzyme consists of 483 residues. The sequence could be easily aligned with the sequence of Escherichia coli serine hydroxymethyltransferase. The primary structural homology between the rabbit and E. coli enzymes is about 42%. The importance of the primary and predicted secondary structural homology between the two enzymes is discussed.

Amino Acid Sequence↗
PubMed1987

A simple microcomputer program for predicting the secondary structure of proteins.

An Applesoft BASIC program is presented which applies the Chou-Fasman and Garnier-Osguthorpe-Robson rules for predicting the secondary structure of proteins. This program plots the result of predictions and shows the charge distribution and the consensus prediction. The program is run on an Apple IIe and the results are printed by an Imagewriter printer.

Computers↗
PubMed1987

PROTEUS: a suite of programs for prediction of structural features of proteins using an Apple IIe.

We have implemented several algorithms, developed by various authors for predicting structural features of proteins from their primary structure, on an Apple IIe and collected them in a suite, named PROTEUS. This suite incorporates: (i) methods for predicting secondary structure; (ii) the algorithm for computing the hydropathy profile using one out of five available sets of parameters; (iii) the algorithms for calculating the hydrophobic moment plot; and (iv) for performing the amphipathic analysis using one out of four available sets of parameters. The suite has a utility program for storing on a disk the sequence to be analysed. As an example, we applied some of the methods included in PROTEUS to predict the structure of a mitochondrial leader peptide. The results suggest the occurrence of structural features possibly related to the import of proteins into mitochondria.

Algorithms↗
PubMed1986

The complete amino acid sequences of cytosolic and mitochondrial aspartate aminotransferases from horse heart, and inferences on evolution of the isoenzymes.

We report here the complete amino acid sequences of the cytosolic and mitochondrial aspartate aminotransferases from horse heart. The two sequences can be aligned so that 48.1% of the amino acid residues are identical. The sequences have been compared with those of the cytosolic isoenzymes from pig and chicken, the mitochondrial isoenzymes from pig, chicken, rat, and human, and the enzyme from Escherichia coli. The results suggest that the mammalian cytosolic and mitochondrial isoenzymes have evolved at equal and constant rates whereas the isoenzymes from chicken may have evolved somewhat more slowly. Based on the rate of evolution of the mammalian isoenzymes, the gene-duplication event that gave rise to cytosolic and mitochondrial aspartate aminotransferases is estimated to have occurred at least 10(9) years ago. The cytosolic and mitochondrial isoenzymes are equally related to the enzyme from E. coli; the prokaryotic and eukaryotic enzymes diverged from one another at least 1.3 X 10(9) years ago.

Amino Acid Sequence↗
PubMed1985

The primary structure of mitochondrial aspartate aminotransferase from human heart.

The complete amino acid sequence of the mitochondrial aspartate aminotransferase (L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1) from human heart has been determined based mainly on analysis of peptides obtained by digestion with trypsin and by chemical cleavage with cyanogen bromide. Comparison of the sequence with those of the isotopic isoenzymes from pig, rat and chicken showed 27, 29 and 55 differences, respectively, out of a total of 401 amino acid residues. Evidence for structural microheterogeneity at position 317 has also been obtained.

Amino Acid Sequence↗