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Biomedical subjects

P Penczek

Publications and source records attributed to P Penczek.

At least 19 recordsLinked to original sources
PubMed1997

Direct localization of the tRNAs within the elongating ribosome by means of neutron scattering (proton-spin contrast-variation).

A new technique for neutron scattering, the proton-spin contrast-variation, improves the signal-to-noise ratio more than one order of magnitude as compared to conventional techniques. The improved signal enables small RNA ligands within a large deuterated ribonucleic acid-protein complex to be measured. We used this technique to determine the positions of the two tRNAs within the elongating ribosome before and after translocation. Using a four-sphere model for each of the L-shaped tRNAs, unequivocal solutions were found for the localization of the mass centre of both tRNAs. The centre of gravity is located in the interface cavity separating the ribosomal subunits near the neck of the 30 S subunit. It moves during translocation by 12(+/-4) A towards the head of the 30 S subunit and slightly towards the L1 protuberance of the 50 S subunit.

Base Sequence↗
PubMed1996

Three-dimensional reconstruction of the Escherichia coli 30 S ribosomal subunit in ice.

Three-dimensional (3D) reconstructions of both the heat-activated and non-activated 30 S subunit of the Escherichia coli 70 S ribosome were obtained from a frozen hydrated specimen preparation at 1/37 A-1 resolution. Well-characterized features that can be identified in both reconstructions are the head, the base, the platform and the cleft formed between the head and the platform. Comparisons between the 3D maps of 30 S subunits at 0 degree C, heat-activated at 37 degrees C, and the 30 S subunit portion identified in the cryo-3D map of 70 S ribosome reveal conformational changes the subunit probably undergoes during inactive-active transition and upon association with the 50 S subunit. These comparisons also allow us to localize the sites of association of 30 S and 50 S subunits.

Escherichia coli↗
PubMed1996

Direct visualization of A-, P-, and E-site transfer RNAs in the Escherichia coli ribosome.

Transfer RNA (tRNA) molecules play a crucial role in protein biosynthesis in all organisms. Their interactions with ribosomes mediate the translation of genetic messages into polypeptides. Three tRNAs bound to the Escherichia coli 70S ribosome were visualized directly with cryoelectron microscopy and three-dimensional reconstruction. The detailed arrangement of A- and P-site tRNAs inferred from this study allows localization of the sites for anticodon interaction and peptide bond formation on the ribosome.

Anticodon↗
PubMed1995

A model of protein synthesis based on cryo-electron microscopy of the E. coli ribosome.

The ribosome is formed by assembly of proteins and nucleic acids, and synthesizes proteins according to genetic instructions in all organisms. Many of the biochemical steps of this fundamental process are known, but a detailed understanding requires a well-defined structural model of the ribosome. Electron microscopy combined with image reconstruction of two-dimensional crystals or single ribosomes has been the most promising technique, but the resolution of the resulting models has been insufficient. Here we report a 25-A reconstruction of the ribosome from Escherichia coli, obtained by combining 4,300 projections of ice-embedded single particles. Our new reconstruction reveals a channel in the small ribosomal subunit and a bifurcating tunnel in the large subunit which may constitute pathways for the incoming message and the nascent polypeptide chain, respectively. Based on these new findings, a three-dimensional model of the basic framework of protein synthesis is presented.

Bacterial Proteins↗
PubMed1995

Automatic particle picking from electron micrographs.

A computer program for automatic particle picking based on textural methods is proposed. The technique relies on the evaluation of certain textural parameters for data windows containing single particles, and those containing undesirable material. These parameters are manipulated by a discriminant analysis routine for determining the rules of classification between the different categories. The effectiveness of the method was demonstrated by application to electron micrographs of 70S Escherichia coli ribosomes.

Algorithms↗
PubMed1995

Double-tilt electron tomography.

Fidelity of tomographic reconstructions is improved and reconstruction artifacts are reduced, without increasing the number of projections, by combining tilt series taken around two orthogonal axes. Test reconstructions were made from high-voltage EM of rat liver mitochondria in a 0.6 micron thick plastic section. A number of schemes for selecting tilt angles for the projections are compared. A new method for aligning fiducial markers is described. It uses an iterative algorithm to determine the shift, scale, in-plane rotation and tilt angle for each tilt image, enforcing agreement of the expected locations of the fiducial markers in 3D space. These 3D locations are used to find the orientation between two tilt series and to merge both sets of projections.

Animals↗
PubMed1994

Quaternary structure of Octopus vulgaris hemocyanin. Three-dimensional reconstruction from frozen-hydrated specimens and intramolecular location of functional units Ove and Ovb.

A frozen-hydrated sample of Octopus vulgaris hemocyanin was imaged at 0 degree and 40 degrees tilt angle under low dose conditions by transmission electron microscopy. A three-dimensional reconstruction by the method of random conical tilt series produced a three-dimensional volume to which a D5 symmetry was applied. Examination of serial sections in the volume and surface representation at various thresholds allowed the five arches containing functional unit Ovg to be localized at the interdimeric subunit groove. In another set of experiments specific polyclonal antibodies were used to label functional units Ovb and Ove in the cylinder wall. The observation of the negatively stained immunocomplexes showed that Ovb is located in the external tiers of functional units and Ove in the internal tier. These results suggest that the direction of the polypeptide chains in the cylinder wall may be only partially antiparallel. A model of the quaternary structure is proposed with the following features: (1) the external tiers of functional units comprise four units each (Ova-d) coming from a single polypeptide chain; (2) the internal tier comprises two functional units from each polypeptide chain (Ove-f); (3) the interdimeric subunit arches connect the two copies of a single functional unit (Ovg) located in each polypeptide chain.

Animals↗
PubMed1994

The internal compartmentation of rat-liver mitochondria: tomographic study using the high-voltage transmission electron microscope.

The three-dimensional organization of the internal compartments of conventionally fixed and embedded rat-liver mitochondria has been determined by tomographic reconstruction from tilt-series images collected on the Albany high-voltage electron microscope. The results indicate that the inner membranes of these organelles are predominantly tubular in the orthodox (expanded matrix) conformation, as previously suggested by scanning electron microscopy. In the condensed (contracted matrix) conformation, the intracristal space opens up into large irregularly shaped compartments which are connected to each other and to the external (intermembrane) space by tubes with approximately the same diameter (20 nm) as those observed in the orthodox state. These results raise several questions, in particular about the nature of the structural transitions that occur in the cristae during matrix expansion and contraction, and about the influence of inner-membrane shape on the diffusion of ions and metabolites between the intracristal and intermembrane compartments.

Animals↗
PubMed1993

Three-dimensional architecture of human alpha 2-macroglobulin transformed with methylamine.

A frozen-hydrated sample embedded in vitreous ice of human alpha 2-macroglobulin transformed by methylamine was imaged by cryoelectron microscopy and reconstructed in three dimensions. In the reconstruction, the cage-like architecture of this protease inhibitor is fully revealed with a clear visualization of two lozenge-shaped lateral walls connected by thin bridges. The shape and dimensions of the internal cavity normally containing the trapped protease(s) is described. The possible locations of the thiol ester sites and inter-subunit connections are also discussed.

Image Processing, Computer-Assisted↗
PubMed1992

Alignment, classification, and three-dimensional reconstruction of single particles embedded in ice.

Cryo-electron microscopy of single biological particles poses new challenges to digital image processing due to the low signal-to-noise ratio of the data. New tools have been devised to deal with important aspects of 3-D reconstruction following the random-conical data collection scheme: (a) a new shift-invariant function has been derived, which promises to facilitate alignment and classification of single particle projections; (b) a new method of orientation search is proposed, which makes it possible to relate random-conical data sets to one another prior to reconstruction; and (c) the foundation is laid for a 3-D variance estimation which utilizes the oversampling of 3-D angular space by projections in the random-conical reconstruction scheme.

Algorithms↗
PubMed1992

Three-dimensional reconstruction of single particles embedded in ice.

Single particles embedded in ice pose new challenges for image processing because of the intrinsically low signal-to-noise ratio of such particles in electron micrographs. We have developed new techniques that address some of these problems and have applied these techniques to electron micrographs of the Escherichia coli ribosome. Data collection and reconstruction follow the protocol of the random-conical technique of Radermacher et al. [J. Microscopy 146 (1987) 113]. A reference-free alignment algorithm has been developed to overcome the propensity of reference-based algorithms to reinforce the reference motif in very noisy situations. In addition, an iterative 3D reconstruction method based on a chi-square minimization constraint has been developed and tested. This algorithm tends to reduce the effects of the missing angular range on the reconstruction, thereby facilitating the merging of random-conical data sets obtained from differently oriented particles.

Algorithms↗
PubMed1991

Three-dimensional reconstruction of the 70S Escherichia coli ribosome in ice: the distribution of ribosomal RNA.

A reconstruction, at 40 A, of the Escherichia coli ribosome imaged by cryo-electron microscopy, obtained from 303 projections by a single-particle method of reconstruction, shows the two subunits with unprecedented clarity. In the interior of the subunits, a complex distribution of higher mass density is recognized, which is attributed to ribosomal RNA. The masses corresponding to the 16S and 23S components are linked in the region of the platform of the small subunit. Thus the topography of the rRNA regions responsible for protein synthesis can be described.

Computer Graphics↗
PubMed1989

Analysis of the multi-channel epileptiform EEG using the Markov chains formalism.

A multi-level scheme of syntactic reduction of the epileptiform EEG data is briefly discussed and the possibilities it opens up in describing the dynamic behaviour of a multi-channel system are indicated. A new algorithm for the inference of a Markov network from finite sets of sample symbol strings is introduced. Formulae for the time-dependent state occupation probabilities, as well as joint probability functions for pairs of channels, are given. An exemplary case of analysis in these terms, taken from an investigation of anticonvulsant drug effects on EEG seizure patterns, is presented.

Algorithms↗
PubMed1987

The use of a multi-channel Kalman filter algorithm in structural analysis of the epileptic EEG.

An extension of the Kalman filter algorithm to the multi-channel case is presented and its application as a segmenting procedure in the analysis of the epileptic EEG is discussed. An analytical example of structural analysis, using the segments extracted by the proposed filter, is presented for a particular set of 4-channel EEG recordings. This analysis is shown to be especially fruitful if the autoregressive coefficients - a byproduct of the filtering procedure - are used to estimate the information flow between the channels by the calculation of partial as well as directed coherences for the representative segments.

Algorithms↗
PubMed1986

Segmentation of the epileptic EEG by means of a finite state automaton.

The aims and merits of the syntactical decomposition of the epileptic EEG are discussed in some detail. General assumptions in the design of segmenting algorithm--an essential part of the reduction of data program--are considered and a working example of segmenting routine is presented. This routine is designed as a finite state automaton, equivalent to a formal regular grammar. The performance of the routine on two sets of experimental data is presented and its intrinsic stability is shown in terms of cluster analysis of the segments produced. Means of extension of the proposed scheme to other cases of potential interest as well as possible applications in the higher level syntactical analysis are pointed out.

Computers↗
PubMed1986

Syntactic analysis of the epileptic electroencephalogram.

A full-scale scheme of the multi-level syntactic analysis of the epileptic EEG is presented and discussed in detail. An exhaustive description of basic procedures necessary to obtain a meaningful description of any set of epileptic EEG recordings is given; these procedures include algorithms for extraction of various elementary peaks from the EEG signal, for segmentation of the recording, for classification of the segments found into distinct groups of characteristic patterns and, finally, for calculation of the Levenshtein distance between the individual recordings, each recording regarded as a string of symbols taken to represent different classes of patterns. An example of global analysis in terms of such a distance, performed on a typical set of epileptic EEG recordings, is supplied to prove the feasibility of the scheme proposed.

Algorithms↗