Showing papers by "David Eisenberg published in 1996"

Assessing the performance of fold recognition methods by means of a comprehensive benchmark.

Abstract: Recently there has been an explosion of methods for fold recognition. These methods seek to align a protein sequence to a three-dimensional structure and measure the compatibility of the sequence to the structure. In this work, we present a benchmark to assess the performance of such methods. The benchmark consists of a set of protein sequences matched by superposition to known structures. This set covers a wide range of protein families, and includes matching proteins with insignificant sequence similarity. To demonstrate the usefulness of this benchmark, we apply it here to compare different fold-recognition methods developed through the years in our group as well as several sequence-sequence substitution matrices. The results show that "global-local" alignments are superior to either local or global alignments. The most effective sequence-sequence matching matrix is the Gonnet table. The best performance overall is obtained by a method which combines the 3D-1D profiles of Bowie et al. with a substitution matrix and takes into account residue pairwise interactions.

Crystal structure of diphtheria toxin bound to nicotinamide adenine dinucleotide.

Abstract: Diphtheria toxin (DT), a 58 kDa protein secreted by lysogenic strains of Corynebacterium diphtheriae, causes the disease diphtheria in humans by gaining entry into the cytoplasm of cells and inhibiting protein synthesis. Specifically, the catalytic (C) domain of DT transfers the ADP-ribose group of NAD to elongation factor-2 (EF-2), rendering EF-2 inactive. In order to investigate how the C-domain of DT binds NAD and catalyzes the ADP-ribosylation of EF-2, the crystal structure of DT in complex with NAD has been determined to 2.3 A resolution. This is the first crystal structure of an ADP-ribosyltransferase (ADP-RT) enzyme in complex with NAD and suggests the features of the ADP-RT fold which are important for NAD binding. The conformation of NAD in the complex and the proximity of the Glu148 carboxylate group of the C-domain to the scissile, N-glycosidic bond of NAD suggest plausible modes of catalysis of the ADP-ribosylation reaction. Residues 39-46 of the active-site loop of the C-domain become disordered upon NAD binding, suggesting a potential role for this loop in the recognition of the ADP-ribose acceptor substrate, EF-2. The negatively charged phosphates and two ribose hydroxyls of NAD are not in direct contact with any atoms of the C-domain. Instead, they form an exposed surface which appears to be presented for recognition by EF-2. Structural alignments of the DT-NAD complex with the structures of other members of the ADP-RT family suggest how NAD may bind to these other enzymes.

A missing link in cupredoxins: crystal structure of cucumber stellacyanin at 1.6 A resolution.

Abstract: Stellacyanins are blue (type I) copper glycoproteins that differ from other members of the cupredoxin family in their spectroscopic and electron transfer properties. Until now, stellacyanins have eluded structure determination. Here we report the three-dimensional crystal structure of the 109 amino acid, non-glycosylated copper binding domain of recombinant cucumber stellacyanin refined to 1.6 A resolution. The crystallographic R-value for all 18,488 reflections (sigma > 0) between 50-1.6 A is 0.195. The overall fold is organized in two beta-sheets, both with four beta-stands. Two alpha-helices are found in loop regions between beta-strands. The beta-sheets form a beta-sandwich similar to those found in other cupredoxins, but some features differ from proteins such as plastocyanin and azurin in that the beta-barrel is more flattened, there is an extra N-terminal alpha-helix, and the copper binding site is much more solvent accessible. The presence of a disulfide bond at the copper binding end of the protein confirms that cucumber stellacyanin has a phytocyanin-like fold. The ligands to copper are two histidines, one cysteine, and one glutamine, the latter replacing the methionine typically found in mononuclear blue copper proteins. The Cu-Gln bond is one of the shortest axial ligand bond distances observed to date in structurally characterized type I copper proteins. The characteristic spectroscopic properties and electron transfer reactivity of stellacyanin, which differ significantly from those of other well-characterized cupredoxins, can be explained by its more exposed copper site, its distinctive amino acid ligand composition, and its nearly tetrahedral ligand geometry. Surface features on the cucumber stellacyanin molecule that could be involved in interactions with putative redox partners are discussed.

Assigning amino acid sequences to 3-dimensional protein folds.

Abstract: With the advent of genome sequencing projects, the amino acid sequences of thousands of proteins are determined every year. Each of these protein sequences must be identified with its function and its 3-dimensional structure for us to gain a full understanding of the molecular biology of organisms. To meet this challenge, new methods are being developed for fold recognition, the computational assignment of newly determined amino acid sequences to 3-dimensional protein structures. These methods start with a library of known 3-dimensional target protein structures. The new probe sequence is then aligned to each target protein structure in the library and the compatibility of the sequence for that structure is scored. If a target structure is found to have a significantly high compatibility score, it is assumed that the probe sequence folds in much the same way as the target structure. The fundamental assumptions of this approach are that many different sequences fold in similar ways and there is a relativel...

Assigning amino acid sequences to 3-dimensional protein folds

Abstract: With the advent of genome sequencing projects, the amino acid sequences of thousands of proteins are determined every year. Each of these protein sequences must be identified with its function and its 3-dimensional structure for us to gain a full understanding of the molecular biology of organisms. To meet this challenge, new methods are being developed for fold recognition, the computational assignment of newly determined amino acid sequences to 3-dimensional protein structures. These methods start with a library of known 3-dimensional target protein structures. The new probe sequence is then aligned to each target protein structure in the library and the compatibility of the sequence for that structure is scored. If a target structure is found to have a significantly high compatibility score, it is assumed that the probe sequence folds in much the same way as the target structure. The fundamental assumptions of this approach are that many different sequences fold in similar ways and there is a relativel...

Unusual trigonal-planar copper configuration revealed in the atomic structure of yeast copper-zinc superoxide dismutase.

Abstract: The three-dimensional structure of yeast copper−zinc superoxide dismutase (CuZnSOD) has been determined in a new crystal form in space group R32 and refined against X-ray diffraction data using difference Fourier and restrained crystallographic refinement techniques. The unexpected result is that the copper ion has moved approximately 1 A from its position in previously reported CuZnSOD models, the copper−imidazolate bridge is broken, and a roughly trigonal planar ligand geometry characteristic of CuI rather than CuII is revealed. Final R values for the two nearly identical room temperature structures are 18.6% for all 19 149 reflections in the 10.0−1.7 A resolution range and 18.2% for 17 682 reflections (F > 2σ) in the 10.0−1.73 A resolution range. A third structure has been determined using X-ray data collected at −180 °C. The final R value for this structure is 19.0% (Rfree = 22.9%) for all 24 356 reflections in the 10.0−1.55 A resolution range. Virtually no change in the positions of the ligands to th...

A Challenging Case for Protein Crystal Structure Determination: the Mating Pheromone Er-1 from Euplotes raikovi

Abstract: Four different phasing methods have been applied to the determination of the crystal structure of the 40 amino-acid mating pheromone of the unicellular ciliated protozoan Euplotes raikovi. The difficulties, failures and successes in attempts to solve the structure by: (1) molecular replacement, (2) direct phasing using the `Shake and Bake' algorithm, (3) isomorphous replacement, and (4) multiple-wavelength anomalous disper- sion are described. The structure was first solved by molecular replacement, and then was the first successful structure determination by `Shake and Bake' without the direct involvement of its authors. A description of the current status of the high-resolution refinement of the structure is also given. The model is refined against 1 A resolution data to an R factor of 12.9%, and includes H atoms and discretely disordered side chains.

Three-dimensional profiles for measuring compatibility of amino acid sequence with three-dimensional structure.

Abstract: Publisher Summary The three-dimensional (3-D) profile method is a general approach for assessing whether a given sequence is compatible with a particular 3-D structure. The method has been used in structure identification, structure verification, de novo protein folding, and secondary structure prediction. This chapter discusses the approaches to fold identification and verification of the correctness of structures using 3-D profiles, three different methods for generating the 3-D profile, and the application of these methods to fold identification and structure verification. One of the uses of 3-D profiles in structure prediction is to determine whether a sequence of unknown structure is compatible with a known 3-D structure. Another application of 3-D profiles is to verify that a proposed model of a protein structure is correct. This is done by testing whether the sequence is compatible with its own structure.

A Structure-Based Model of Diphtheria Toxin Action

Abstract: Scientific efforts to find the cause of the disease diphtheria date back to at least 1740, when the clergyman Jonathan Dickenson, later the first president of Princeton, wrote his paper “Observations on that terrible Disease vulgarly called the Throat Distemper with advices as to the Method of cure.”1 Dickenson reported his observations during the diphtheria epidemic that swept the northern American colonies in the years 1735–1740, eventually infecting in attenuated form about a fourth of the people in Boston. He wrote: “The first Assault was in a Family about ten Miles from me, which proved fatal to eight of the Children in about a Fortnight. Being called to visit the distressed Family, I found upon my arrival, one of the Children newly dead, which gave me the Advantage of a Dissection, and thereby a better Acquaintance with the Nature of the Disease, than I could otherwise have had.... It frequently begins with a slight Indisposition, much resembling an ordinary Cold, with a listless Habit, a slow and scarce discernable Fever, some soreness of the Throat and Tumefaction of the Tonsils: and perhaps a running of the Nose, the Countenance pale, and the eyes dull and heavy. The patient is not confined, nor any Danger apprehended for some Days, till the Fever gradually increases, the whole Throat, and sometimes the Roof of the Mouth and Nostrils are covered with a cankerous Crust.... When the lungs are thus affected, the Patient is first afflicted with a dry hollow Cough, which is quickly succeeded with an extraordinary Hoarseness and total Loss of the Voice, with the most distressing asthmatic Symptoms and difficulty of Breathing, under which the poor miserable creature struggles, until released by a perfect Suffocation, or Stoppage of the Breath.”