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S Marco

Publications and source records attributed to S Marco.

34 records · Page 2Linked to original sources
PubMed1997

Symmetric GroEL-GroES complexes can contain substrate simultaneously in both GroEL rings.

Incubation of rhodanese with hche aperonins GroEL and GroES (1:2 GroEL14:GroES7 molar ratio) under functional and steady state conditions for ATP leads to the formation of a high proportion of rhodanese-bound symmetric complexes (GroEL14(GroES7)2), as revealed by native electrophoresis. Aliquots of such samples were observed under the electron microscope, and the symmetric particles were classified using neuronal networks and multivariate statistical analysis. Three different populations of symmetric particles were obtained which contained substrate in none, one or both GroEL cavities, respectively. The presence of substrate in the symmetric complexes under functional conditions supports their role as active intermediates in the protein folding cycle. These results also suggest that symmetric GroEL-GroES complexes can use both rings simultaneously for folding, probably increasing the efficiency of the reaction.

Adenosine Triphosphate↗
PubMed1997

Conformational changes in the GroEL oligomer during the functional cycle.

The conformational changes that the GroEL oligomer undergoes upon nucleotide and cochaperonin GroES binding have been studied using electron microscopy and image processing techniques. Average side views of the three allosteric states (TT, TR, and RR, which correspond to none, one, or both of the two heptameric rings of the GroEL oligomer occupied by nucleotide, respectively) of GroEL and GroEL-GroES complexes for ADP, ATP, and two nonhydrolyzable analogs (AMP-PNP and ATP gamma S) have been obtained at 20-25 A resolution. Both AMP-PNP and ATP induce similar conformational shifts in the apical domains of GroEL. At the TR state, only one of the GroEL rings shows an upward and outward movement of the apical domains ("open state"). At the RR state for AMP-PNP and ATP, both GroEL rings undergo conformational changes, albeit of different magnitude, giving rise to a structurally asymmetric particle (one ring in the "open" state, while the other is in an "intermediate" state). These changes are also observed when GroEL is incubated with ADP and Pi, but not with ADP, which suggests that upon ATP binding, GroEL undergoes a conformational change that is partly maintained after ATP hydrolysis and as long as ADP and Pi are bound to the GroEL ring. The conformational changes undergone by GroEL are discussed within the framework of a proposed GroEL cycle mechanism.

Adenine Nucleotides↗
PubMed1997

Paget's disease.

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Alleles↗
PubMed1996

Xmipp: An Image Processing Package for Electron Microscopy

We present in this article a specialized suite of image processing programs primarily aimed at obtaining the three-dimensional reconstruction of biological specimens from large sets of projection images obtained by transmission electron microscopy. The programs are written in ANSI-C and use X-Windows for graphical output. A number of parallel extensions using PVM and PARMACS are provided. The programs are freely available by anonymous ftp at ftp.cnb.uam.es.

Journal Article↗
PubMed1994

The formation of symmetrical GroEL-GroES complexes in the presence of ATP.

The incubation of chaperonins cpn60 (GroEL) and cpn10 (GroES) from E. coli in the presence of Mg-ATP and KCl generates the formation, as revealed by electron microscopy, of GroEL-GroES complexes with a symmetrical shape in which one toroidal GroES oligomer is bound to each end of the tetradecameric GroEL aggregate (1:2 GroEL:GroES oligomer molar ratio). The symmetrical complexes are not observed in the presence of ADP or the non-hydrolyzable ATP analog, ATP gamma S, where only asymmetrical complexes (1:1 GroEL:GroES oligomer molar ratio) are formed. These results suggest that ATP hydrolysis is required for the formation of symmetrical complexes.

Adenosine Triphosphate↗
PubMed1994

The molecular chaperone TF55. Assessment of symmetry.

TF55-like factor from Sulfolobus solfataricus was purified to homogeneity and analyzed by electron microscopy and image analysis to determine the symmetries of these particles. Three different procedures were used to analyze the electron micrographs: (1) fuzzy-set based classification of the particles according to their rotational power spectra; (2) multivariate statistical analysis based on singular value decomposition; (3) circular harmonic analysis. Averages obtained from the three methods show unequivocally that the TF55-like complex presents a 9-fold symmetry.

Archaeal Proteins↗
PubMed1994

Reversible interaction of beta-actin along the channel of the TCP-1 cytoplasmic chaperonin.

The cytoplasm of eukaryotes contains a heteromeric toroidal chaperonin assembled from the t-complex TCP-1 and several other related polypeptides. The structure of the TCP-1 cytoplasmic chaperonin and that of the binary complex formed between this chaperonin and unfolded beta-actin have been studied using electron microscopy and image processing techniques. Two-dimensional averaging of front views reveals a circular stain-excluding mass surrounding a central stain-penetrating region in which the stain is excluded upon actin binding. Sections of a three-dimensional reconstruction of the chaperonin show that the inner core is an empty channel that becomes filled upon binary complex formation with unfolded beta-actin. Upon incubation with Mg-ATP, the beta-actin:chaperonin complex discharges the actin such that the chaperonin central cavity reappears. Side views from different forms of TCP-1 reveals that upon Mg-ATP binding, the cytoplasmic chaperonin undergoes a structural rearrangement that is confirmed using a new classification method.

Actins↗
PubMed1994

Compliance with pharmacologic prophylaxis and therapy in bronchial asthma.

Between January 1 and December 31, 1988, 288 children [185 boys and 103 girls, mean age 8.75 +/- 4.98 years (range 2.50 to 16.83)], followed at the Outpatient Clinic for Lung Diseases of the University of Milan 5th Pediatric Department, were interviewed blindly in order to assess their compliance with pharmacologic therapy. All children were suffering from episodic, frequent, or chronic asthma requiring therapy as needed and preventive drugs for at least 30 days. Prophylaxis (including cromolyn, beclomethasone, theophylline retard, ketotifen, oxatomide, albuterol, and prednisone, alone or in combinations) and therapy in case of symptoms (albuterol, with or without theophylline prompt or beclomethasone) were prescribed. A study questionnaire was completed 30 to 45 days after the prescription with no advance warning by a physician unaware of the prophylactic and therapeutic prescriptions. Out of the 288 patients, 31 (10.8%) failed to return for the second visit. Understanding of prophylaxis was full in 61.1% of cases, partial in 23.3% and nil in 4.9%. Therapy as needed was fully remembered by 77.1% of parents, partially by 9.4% and totally forgotten by 2.8%. Compliance with single-drug prophylaxis was full in 60.1% of cases, partial in 22.2% and nil in 6.4%. Statistical analysis showed compliance was significantly better for ketotifen than for disodium cromoglycate (chi squared 9.85, P < .02), for ketotifen than for theophylline (chi squared 9.98, P < .02), and for beclomethasone than for theophylline (chi squared 8.77, P < .05).(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Inhalation↗
PubMed1993

A structural model for the GroEL chaperonin.

Individual particle analysis of end views from negatively stained specimens of purified GroEL from Escherichia coli showed the presence of two different particle populations, those with a six-fold symmetry and those with a seven-fold symmetry, when studied at pH 7.7 and 5.0. Image processing of particles from frozen-hydrated specimens revealed at both pH values a homogeneous population of particles with a strong seven-fold symmetry component and an average image with seven asymmetric units. Biochemical analysis of purified GroEL showed unequivocally the presence of a single polypeptide with the N-terminal sequence identical to that of GroEL. These results are compatible with a structural model of GroEL as an asymmetric aggregate built up by two rings of seven-fold and six-fold symmetries, respectively.

Amino Acid Sequence↗
PubMed1992

Streptomyces lividans possesses a GroEL-like chaperonin.

Streptomyces lividans grown at 45 degrees C produces a GroEL-like chaperonin. This protein is specifically synthesized in bacterial cell cultures upon heat shock induction. It has a similar size (62 kDa) to the GroEL-like proteins from Escherichia coli and Bacillus subtilus and shows immunological cross-reaction with serum raised against GroEL from E. coli. The S. lividans 62-kDa protein assembles into oligomers around 20S that show a morphology consistent with a barrel showing six-fold and seven-fold symmetries as previously described in E. coli and B. subtilis.

Bacterial Proteins↗
PubMed1992

Three-dimensional structure of T3 connector purified from overexpressing bacteria.

The bacteriophage T3 connector has been purified from overexpressed protein in Escherichia coli, harboring a plasmid containing the gene encoding p8 protein. The connector, which is composed of 12 copies of p8, has been crystallized in two-dimensional sheets and studied by electron microscopy from negatively stained specimens. A two-dimensional Fourier filtering and averaging procedure was performed with crystalline specimens. In addition, single particle averaging techniques were used with other preparations. The average images obtained from these two approaches gave similar results. A three-dimensional reconstruction from two-dimensional crystals of T3 connectors was obtained by collecting several sets of tilted views and using standard Fourier procedures. The resolution of the three-dimensional map was 1.65 nm. The reconstructed connector shows two main domains: a wider one with 12 small units in the periphery and with an external diameter of 14.9 nm, and a smaller one with 8.5 nm diameter. The height of the reconstructed connector has been determined to be around 8.5 nm. The reconstruction clearly shows an internal open channel running along the longitudinal axis of the particle and having an average diameter of 3.7 nm.

Capsid↗
PubMed1991

Electron microscopy study of GroEL chaperonin: different views of the aggregate appear as a function of cell growth temperature.

We have studied two members of the family of morphogenetic factors or chaperonins, the GroEL-like factors from Escherichia coli and Bacillus subtilis, in order to determine the possible structural basis of their related function in promoting the correct and efficient assembly of biological oligomers. The main objective of this work has been to study by transmission electron microscopy the possible changes that these factors may undergo when subjected to a number of different conditions such as changes in temperature in vivo and in pH in vitro. We applied both rotational and multivariate statistical analyses of single particles to images of GroEl-like aggregates from the two bacteria. The most striking result is the finding of two distinct "front views" of these aggregates, from both E. coli and B. subtilis. One view, which has not been described earlier, shows a sixfold symmetry and is most abundant at growing temperatures below 37 degrees C. After heat shock, a view showing seven morphological units becomes dominant. On the basis of our analysis it is clear that GroEL-like morphogenetic factors from two unrelated bacteria such as E. coli and B. subtilis present two distinct views: one sixfold and the other sevenfold. Their relative percentage of appearance is related to the temperature at which the cells were grown and also to the storage conditions (pH).

Bacillus subtilis↗
PubMedDate not supplied

Three-dimensional reconstruction of the sevenfolded form of Bacillus subtilis Gro EL Chaperonin.

Bacillus subtilis grown at 42 degrees C produces a major form of Gro EL-like chaperonin that has been analyzed by electron microscopy. Most of the views show a clear sevenfold symmetry when studied by rotational analysis. The particles were classified into defined families by multivariate analysis and supervised fuzzy-set classification methods, and those belonging to a sevenfold family were averaged to produce a two-dimensional representative projection. These selected particles were then used, when titled by 55 degrees in the microscope goniometer stage, as the starting projections for a three-dimensional reconstruction protocol based on the random conical tilt series method. The resulting reconstruction shows the Gro EL-like chaperonin from B. subtilis as a cylindrical body with seven well defined lobules arranged almost parallel to the longitudinal axis of the particle. There is a channel that is placed along this axis and appears fully open in both sides. The geometry of the channel is polar and presents differences in both faces of the particle.

Bacillus subtilis↗
PubMedDate not supplied

Current concepts on myocardial depressant factor.

In the present work the authors have tried to offer a vast and detailed summary of theories and questions concerning the role of the MDF in shock. One of the major problems that surrounds this molecule is the myocardial contractility depression, the solution of which could allow a more rationalistic therapeutic approach to that which remains one of the most complex and delicate clinical framework.

Animals↗