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M Levitt

Publications and source records attributed to M Levitt.

At least 37 records · Page 2Linked to original sources
PubMed1999

Structure-based conformational preferences of amino acids.

Proteins can be very tolerant to amino acid substitution, even within their core. Understanding the factors responsible for this behavior is of critical importance for protein engineering and design. Mutations in proteins have been quantified in terms of the changes in stability they induce. For example, guest residues in specific secondary structures have been used as probes of conformational preferences of amino acids, yielding propensity scales. Predicting these amino acid propensities would be a good test of any new potential energy functions used to mimic protein stability. We have recently developed a protein design procedure that optimizes whole sequences for a given target conformation based on the knowledge of the template backbone and on a semiempirical potential energy function. This energy function is purely physical, including steric interactions based on a Lennard-Jones potential, electrostatics based on a Coulomb potential, and hydrophobicity in the form of an environment free energy based on accessible surface area and interatomic contact areas. Sequences designed by this procedure for 10 different proteins were analyzed to extract conformational preferences for amino acids. The resulting structure-based propensity scales show significant agreements with experimental propensity scale values, both for alpha-helices and beta-sheets. These results indicate that amino acid conformational preferences are a natural consequence of the potential energy we use. This confirms the accuracy of our potential and indicates that such preferences should not be added as a design criterion.

Amino Acids↗
PubMed1999

Hierarchy of structure loss in MD simulations of src SH3 domain unfolding.

To complement experimental studies of the src SH3 domain folding, we studied 30 independent, high-temperature, molecular dynamics simulations of src SH3 domain unfolding. These trajectories were observed to differ widely from each other. Thus, rather than analyzing individual trajectories, we sought to identify the recurrent features of the high-temperature unfolding process. The conformations from all simulations were combined and then divided into groups based on the number of native contacts. Average occupancies of each side-chain hydrophobic contact and hydrogen bond in the protein were then determined. In the symmetric funnel limit, the occupancies of all contacts should decrease in concert with the loss in total number of native contacts. If there is a lack of symmetry or hierarchy to the unfolding process, the occupancies of some contacts should decrease more slowly, and others more rapidly. Despite the heterogeneity of the individual trajectories, the ensemble averaging revealed an order to the unfolding process: contacts between the N and C-terminal strands are the first to disappear, whereas contacts within the distal beta-hairpin and a hydrogen-bonding network involving the distal loop beta-turn and the diverging turn persist well after the majority of the native contacts are lost. This hierarchy of events resembles but is somewhat less pronounced than that observed in our experimental studies of the folding of src SH3 domain.

Computer Simulation↗
PubMed1999

The PRESAGE database for structural genomics.

The PRESAGE database is a collaborative resource for structural genomics. It provides a database of proteins to which researchers add annotations indicating current experimental status, structural predictions and suggestions. The database is intended to enhance communication among structural genomics researchers and aid dissemination of their results. The PRESAGE database may be accessed at http://presage.stanford.edu/

Databases, Factual↗
PubMed1999

Ab initio protein structure prediction using a combined hierarchical approach.

As part of the third Critical Assessment of Structure Prediction meeting (CASP3), we predict the three-dimensional structures for 13 proteins using a hierarchical approach. First, all possible compact conformations of a protein sequence are enumerated using a highly simplified tetrahedral lattice model. We select a large subset of these conformations using a lattice-based scoring function and build detailed all-atom models incorporating predicted secondary structure. A combined all-atom knowledge-based scoring function is then used to select three smaller subsets from these all-atom models. Finally, a consensus-based distance geometry procedure is used to generate the best conformations from each of the all-atom subsets. With this method, we are able to predict the global topology/shape for all or a large part of the sequence for six out of the thirteen proteins. For two other proteins, the topology/shape for shorter fragments are predicted. This represents a marked improvement in ab initio prediction since CASP was first instigated in 1994.

Algorithms↗
PubMed1999

Treatment of chronic anal fissure with topical glyceryl trinitrate.

PURPOSE: There has been an emerging trend to treat chronic anal fissures by pharmacologic means. In particular, by the topical application of glyceryl trinitrate. However, such treatment has failed to gain wide acceptance within our local surgical community. The aim of this study was to evaluate the efficacy and patient acceptance of topical glyceryl trinitrate in the management of chronic anal fissures. METHODS: A retrospective review was conducted on a consecutive series of 31 patients who were diagnosed as having a chronic anal fissures and were managed by the topical application of 0.2 percent glyceryl trinitrate to the perianal region twice a day. All patients were commenced on a high-fiber diet, and they were reviewed every two weeks until healed or until surgery was required. Patients were subsequently contacted by a mailed questionnaire to determine compliance with therapy, resolution of symptoms, recurrence of symptoms after cessation of therapy, and side effects. RESULTS: Of 31 patients, 27 completed the questionnaire. Only 67 percent (18/27) of patients complied with the treatment as prescribed, and just 56 percent (15/27) achieved complete resolution of their symptoms after three to ten weeks of therapy. Recurrence of symptoms developed in 27 percent (4/15) of these patients after a median follow-up of six months. Side effects were reported in 78 percent (21/27) of patients, and in ten percent (2/21) they were so severe that the patients ceased therapy. CONCLUSION: In this study the treatment of chronic anal fissures with topical glyceryl trinitrate appeared less efficacious than previously reported. In addition, the study highlighted problems with patient compliance, healing rates, and side effects that are associated with this modality therapy.

Administration, Topical↗
PubMed1999

Genetic evolutionary staging of early non-small cell lung cancer: the P53 --> HER-2/NEU --> ras sequence.

INTRODUCTION: The sequence of genetic evolutionary abnormalities that have occurred in a given lung cancer tumor before tumor sampling can be inferred from patterns of intracellular co-occurrence of these abnormalities in tumor cell subpopulations at the time of sampling. The same evolutionary sequences that are present within each lung cancer were evident in intertumor comparisons. METHODS: Correlated cell by cell measurements of cell DNA content, p53, Her-2/neu, and ras proteins were obtained by multiparameter flow cytometry on 46 surgically resected stage I-III primary non-small cell lung cancers. Early evolutionary changes were identified by the fact that they could appear alone in individual cells. Later appearing abnormalities were identified by the fact that they were accompanied by early abnormalities in the same cells. Patients were followed prospectively. Evolutionary patterns observed in individual tumors were correlated with subsequent clinical outcome of patients undergoing surgical resection. RESULTS: Three common patterns were identified: (I) a diploid DNA pathway consisting of the sequence p53 overexpression --> Her-2/neu overexpression --> ras overexpression, (II) an aneuploid DNA pathway with the same p53 --> Her-2/neu --> ras sequence, and (III) a pathway in which none of the intracellular protein measurements made here were abnormal. Fourteen tumors recurred after 11.5 months' median study time. Nine of 12 recurrences in pathways I and II occurred in patients whose tumors were far advanced along these molecular genetic pathways. CONCLUSIONS: Multiparameter cell-based genetic evolutionary studies may be a promising approach for identifying patients with stage I-III non-small cell lung cancer at high risk for recurrence.

Adult↗
PubMed1999

Pre-operative chemoradiotherapy in locally advanced rectal cancer.

BACKGROUND: The aim of the present study was to investigate the effectiveness and toxicity of pre-operative chemoradiation in locally advanced rectal cancer (T3-T4). METHODS: Forty-seven patients were assessed (38 T3 and nine T4 tumours). Pre-operative pelvic radiotherapy was delivered in four fields, 45 Gy in 25 fractions over 5 weeks. Bolus 5-fluorouracil (5-FU) was delivered 500 mg/m2 on days 1, 2, 3 and days 22, 23, 24. Total mesorectal excision of the rectal tumour either by anterior or abdomino-perineal resection was planned at 4-6 weeks from completion of pre-operative treatment. Response to therapy was assessed by fresh macroscopic measurement of the surgical specimen. RESULTS: All patients undergoing chemoradiation completed therapy as planned, with no treatment-related interruptions. The regimen had a low acute toxicity profile with an estimated 50% or greater response in 38 out of 47 patients (four patients had complete responses). Forty-three (97%) of 44 patients who underwent surgery were operable. Patients who were operated on between 4 and 7 weeks had a statistically better response then those who were operated on after 7 weeks (P = 0.013; Fisher's exact test). Eight of 10 patients who were considered to be inoperable prior to the treatment underwent total mesorectal excision with negative radial margins. Anastomotic leakage occurred in four patients (9%); one required surgical intervention. Wound infection occurred in three patients (6%); one patient required re-exploration for haemorrhage. Delayed complications occurred in three patients (6%); one requiring surgery for a stomal stricture. After a median follow-up of 20 months, two patients (4%) had developed local recurrence. CONCLUSION: The pre-operative chemoradiation regimen employed had a low acute toxicity profile and all patients completed therapy. The majority of patients considered inoperable prior to receiving this treatment underwent successful excision. Appropriately fractionated pre-operative chemoradiotherapy is a reasonable option in this disease and deserves further evaluation.

Adenocarcinoma↗
PubMed1999

Pre-operative chemo-radiotherapy in locally advanced rectal cancer: is this the way of future management?

The purpose of the present paper was to update a prospective analysis (H Elsaleh et al. unpubl. data, 1997) investigating the effectiveness and toxicity of pre-operative pelvic radiotherapy with modest dose 5-fluorouracil (5-FU) in locally advanced rectal cancer (T3-T4). A total of 31 patients were assessed (28 T3 and three T4 tumours). Pre-operative pelvic radiotherapy was delivered in four fields, 45 Gy to the International Commission on Radiation Units and Measurements (ICRU) point in 25 fractions over 5 weeks. A radiosensitizing dose of 5-FU was delivered at 500 mg/m2 on days 1, 2 and 3, and days 22, 23 and 24. Mesorectal excision of the rectal tumour either by anterior or abdomino-perineal resection was planned at 4-6 weeks from completion of pre-operative treatment. Response to therapy was assessed by fresh macroscopic measurement of the surgical specimen. Patients had a low toxicity profile; an estimated 50% or greater response was seen in 24 out of 31 (two complete responses). There were no surgical difficulties achieving resection. No late complications were documented, although follow-up was short. In locally advanced rectal cancer, pre-operative chemo-radiotherapy had a low toxicity profile. Appropriately fractionated pre-operative chemo-radiotherapy is a reasonable option in this disease and should be further evaluated. The optimal method of delivery of the radiosensitizing agent (5-FU) is the subject of further investigation.

Adult↗
PubMed1999

A combined approach for ab initio construction of low resolution protein tertiary structures from sequence.

An approach to construct low resolution models of protein structure from sequence information using a combination of different methodologies is described. All possible compact self-avoiding C alpha conformations (approximately 10 million) of a small protein chain were exhaustively enumerated on a tetrahedral lattice. The best scoring 10,000 conformations were selected using a lattice-based scoring function. All-atom structures were then generated by fitting an off-lattice four-state phi/psi model to the lattice conformations, using idealised helix and sheet values based on predicted secondary structure. The all-atom conformations were minimised using ENCAD and scored using a second hybrid scoring function. The best scoring 50, 100, and 500 conformations were input to a consensus-based distance geometry routine that used constraints from each the conformation sets and produced a single structure for each set (total of three). Secondary structures were again fitted to the three structures, and the resulting structures were minimised and scored. The lowest scoring conformation was taken to be the "correct" answer. The results of application of this method to twelve proteins are presented.

Amino Acid Sequence↗
PubMed1999

Differentiation of mouth versus gut as site of origin of odoriferous breath gases after garlic ingestion.

Utilizing the sulfur-containing gases of garlic as probes, we investigated the gut versus mouth origin of odoriferous breath gases. Five individuals ingested 6 g of garlic, and sulfur gases in mouth, alveolar air, and urine samples were measured. The mouth normally contained low concentrations of hydrogen sulfide, methanethiol, and dimethyl sulfide. Immediately after garlic ingestion, transient high concentrations of methanethiol and allyl mercaptan and lesser concentrations of allyl methyl sulfide (AMS), allyl methyl disulfide, and allyl disulfide were observed. With the exception of AMS, all gases were present in far greater concentrations in mouth than alveolar air, indicating an oral origin. Only AMS was of gut origin as evidenced by similar partial pressures in mouth, alveolar air, and urine. After 3 h, AMS was the predominant breath sulfur gas. The unique derivation of AMS from the gut is attributable to the lack of gut and liver metabolism of this gas versus the rapid metabolism of the other gases. Breath odor after garlic ingestion initially originates from the mouth and subsequently from the gut.

Adult↗
PubMed1998

Accuracy of side-chain prediction upon near-native protein backbones generated by Ab initio folding methods.

The ab initio folding problem can be divided into two sequential tasks of approximately equal computational complexity: the generation of native-like backbone folds and the positioning of side chains upon these backbones. The prediction of side-chain conformation in this context is challenging, because at best only the near-native global fold of the protein is known. To test the effect of displacements in the protein backbones on side-chain prediction for folds generated ab initio, sets of near-native backbones (< or = 4 A C alpha RMS error) for four small proteins were generated by two methods. The steric environment surrounding each residue was probed by placing the side chains in the native conformation on each of these decoys, followed by torsion-space optimization to remove steric clashes on a rigid backbone. We observe that on average 40% of the chi1 angles were displaced by 40 degrees or more, effectively setting the limits in accuracy for side-chain modeling under these conditions. Three different algorithms were subsequently used for prediction of side-chain conformation. The average prediction accuracy for the three methods was remarkably similar: 49% to 51% of the chi1 angles were predicted correctly overall (33% to 36% of the chi1+2 angles). Interestingly, when the inter-side-chain interactions were disregarded, the mean accuracy increased. A consensus approach is described, in which side-chain conformations are defined based on the most frequently predicted chi angles for a given method upon each set of near-native backbones. We find that consensus modeling, which de facto includes backbone flexibility, improves side-chain prediction: chi1 accuracy improved to 51-54% (36-42% of chi1+2). Implications of a consensus method for ab initio protein structure prediction are discussed.

Models, Chemical↗
PubMed1998

A unified statistical framework for sequence comparison and structure comparison.

We present an approach for assessing the significance of sequence and structure comparisons by using nearly identical statistical formalisms for both sequence and structure. Doing so involves an all-vs.-all comparison of protein domains [taken here from the Structural Classification of Proteins (scop) database] and then fitting a simple distribution function to the observed scores. By using this distribution, we can attach a statistical significance to each comparison score in the form of a P value, the probability that a better score would occur by chance. As expected, we find that the scores for sequence matching follow an extreme-value distribution. The agreement, moreover, between the P values that we derive from this distribution and those reported by standard programs (e.g., BLAST and FASTA validates our approach. Structure comparison scores also follow an extreme-value distribution when the statistics are expressed in terms of a structural alignment score (essentially the sum of reciprocated distances between aligned atoms minus gap penalties). We find that the traditional metric of structural similarity, the rms deviation in atom positions after fitting aligned atoms, follows a different distribution of scores and does not perform as well as the structural alignment score. Comparison of the sequence and structure statistics for pairs of proteins known to be related distantly shows that structural comparison is able to detect approximately twice as many distant relationships as sequence comparison at the same error rate. The comparison also indicates that there are very few pairs with significant similarity in terms of sequence but not structure whereas many pairs have significant similarity in terms of structure but not sequence.

Animals↗
PubMed1998

Comprehensive assessment of automatic structural alignment against a manual standard, the scop classification of proteins.

We apply a simple method for aligning protein sequences on the basis of a 3D structure, on a large scale, to the proteins in the scop classification of fold families. This allows us to assess, understand, and improve our automatic method against an objective, manually derived standard, a type of comprehensive evaluation that has not yet been possible for other structural alignment algorithms. Our basic approach directly matches the backbones of two structures, using repeated cycles of dynamic programming and least-squares fitting to determine an alignment minimizing coordinate difference. Because of simplicity, our method can be readily modified to take into account additional features of protein structure such as the orientation of side chains or the location-dependent cost of opening a gap. Our basic method, augmented by such modifications, can find reasonable alignments for all but 1.5% of the known structural similarities in scop, i.e., all but 32 of the 2,107 superfamily pairs. We discuss the specific protein structural features that make these 32 pairs so difficult to align and show how our procedure effectively partitions the relationships in scop into different categories, depending on what aspects of protein structure are involved (e.g., depending on whether or not consideration of side-chain orientation is necessary for proper alignment). We also show how our pairwise alignment procedure can be extended to generate a multiple alignment for a group of related structures. We have compared these alignments in detail with corresponding manual ones culled from the literature. We find good agreement (to within 95% for the core regions), and detailed comparison highlights how particular protein structural features (such as certain strands) are problematical to align, giving somewhat ambiguous results. With these improvements and systematic tests, our procedure should be useful for the development of scop and the future classification of protein folds.

Amino Acid Sequence↗
PubMed1998

Complications requiring reoperation following pancreatectomy.

CONCLUSIONS: In this series, the overall reoperative rate following pancreatic surgery is 9%. Complications following pancreatectomy that require reoperation fall into four categories: hemorrhage, infectious, delayed gastric emptying, and anastomotic leak. A delay in the management of these types of complications can be fatal. BACKGROUND: Despite the improvement in the morbidity and mortality rates associated with pancreatic resection, complication still arise that require surgical intervention. This study reviews the pancreatic surgical experience at a major medical center to determine the overall reoperative complication rate. STUDY DESIGN: From 1985 to 1995, 107 patients underwent pancreatic resection. There were 50 pancreaticoduodenectomies, 20 total pancreatectomies, and 37 distal pancreatectomies for 102 periampullary or pancreatic cancers and five for chronic pancreatitis. The operative mortality rate was 6.5% and the morbidity rate was 43%. Ten patients (9%) developed complications that required reoperation. RESULTS: Re-exploration was performed in five patients for hemorrhage. Four patients had bleeding intra-abdominally and one had a suture line bleed. One patient developed a wound infection and fascial necrosis which necessitated reoperation. Three patients were explored for sepsis and one was found to have a pancreatic leak. One patient had persistent gastric outlet obstruction and he required conversion of the gastrojejunostomy to a Roux-en-y anastomosis. The mortality rate for re-exploration was 3/10 (30%).

Humans↗