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We report a comparison of the steady-state absorption and fluorescence spectra of three representative uracil derivatives (uracil, thymine and 5-fluorouracil) in alcoholic solutions. The present results are compared with those from our previous experimental and computational studies of the same compounds in water and acetonitrile. The effects of solvent polarity and hydrogen bonding on the spectra are discussed in the light of theoretical predictions. This comparative analysis provides a more complete picture of the solvent effects on the absorption and fluorescence properties of pyrimidine nucleobases, with special emphasis on the mechanism of the excited state deactivation.

The authors present a new method for the computation of vibrationally resolved optical spectra of large molecules, including the Duschinsky [Acta Physicochim. URSS 7, 551 (1937)] rotation of the normal modes. The method automatically selects the relevant vibronic contributions to the spectrum, independent of their frequency, and it is able to provide fully converged spectra with a quite modest computational time, both in vacuo and in condensed phase. Starting from the rigorous time-dependent expression they discuss indeed in which limits the spectrum of a molecule embedded in a solvent, described as a polarizable continuum, can be computed in a time-independent formalism, defining both nonequilibrium and equilibrium limits. In these cases the polarizable continuum model provides a suitable description of the solvent field. By computing the absorption spectra of anthracene in gas phase and of coumarin C153 in gas phase and cyclohexane, and the phosphorescence spectrum of the unsubstituted coumarin in ethanol they show that the method is fast and efficient.

We demonstrate here that tetracycline (TC) can strongly interact (KD' = 189 +/- 7 nM) with model peptides derived from the C-terminal globular domain of the prion protein, hPrP [173-195], and that interaction concerns residues within the C-terminal half of the helix 2, a short region previously indicated as endowed with ambivalent conformational behavior and implicated in PrP conversion to the beta-sheet-rich, infective scrapie variant. Data have been confirmed by binding studies with the N-terminal truncated 180-195 variant that displays a dissociation constant of 483 +/- 30 nM. Remarkably, TC does not influence the structure of the N-terminally fluoresceinated peptides that both show alpha-helical conformations. Docking calculations and molecular dynamics simulations suggest a direct, strong interaction of the antibiotic with exposed side chain functional groups of threonines 190-193 on the solvent-exposed surface of helix 2., (2006 Wiley-Liss, Inc.)

The first comprehensive quantum mechanical study of solvent effects on the behavior of the two lowest energy excited states of uracil derivatives is presented. The absorption and emission spectra of uracil and 5-fluorouracil in acetonitrile and aqueous solution have been computed at the time-dependent density-functional theory level, using the polarizable continuum model (PCM) to take into account bulk solvent effects. The computed spectra and the solvent shifts provided by our method are close to their experimental counterpart. The S0/S1 conical intersection, located in the presence of hydrogen-bonded solvent molecules by CASSCF (8/8) calculations, indicates that the mechanism of ground-state recovery, involving out-of-plane motion of the 5 substituent, does not depend on the nature of the solvent. Extensive explorations of the excited-state surfaces in the Franck-Condon (FC) region show that solvent can modulate the accessibility of an additional decay channel, involving a dark n/pi* excited state. This finding provides the first unifying explanation for the experimental trend of 5-fluorouracil excited-state lifetime in different solvents. The microscopic mechanisms underlying solvent effects on the excited-state behavior of nucleobases are discussed.

An effective state specific (SS) model for the inclusion of solvent effects in time dependent density functional theory (TD-DFT) computations of excited electronic states has been developed and coded in the framework of the so-called polarizable continuum model (PCM). Different relaxation time regimes can be treated thus giving access to a number of different spectroscopic properties together with solvent relaxation energies of paramount relevance in electron transfer processes. SS and conventional linear response (LR) models have been compared for two benchmark systems (coumarin 153 and formaldehyde in different solvents) and in the limiting simple case of a dipolar solute embedded in a spherical cavity. The results point out the complementarity of LR and SS approaches and the advantages of the latter model especially for polar solvents. The favorable scaling properties of PCM-TD-DFT models in both SS and LR variants and their availability in effective quantum mechanical codes pave the route for the computation of reliable spectroscopic properties of large molecules of technological and/or biological interest in their natural environments.

Recently, optimal control of a photoisomerization reaction in the liquid phase was demonstrated for the first time on the system 3,3(')-diethyl-2,2(')-thiacyanine (NK88). Additionally, the class of cyanines to which the molecule NK88 belongs draws a lot of attention in different recent theoretical publications. Therefore, a better understanding of the molecular dynamics of this molecular system is of special interest. Experiments using the femtosecond pump-supercontinuum probe technique with an excitation wavelength of 400 nm and a spectral range from 370 to 620 nm for the probe beam have been performed. In order to analyze the dynamics properly the time window has been chosen to comprise the characteristic times of the contributing processes, additionally we have employed two solvents, methanol and ethylene glycol, and have conducted anisotropy measurements. The spectroscopic data have been assigned to different molecular states with the help of density functional theory and second-order Moller-Plesset perturbation theory calculations. The analysis of the data has revealed in the most likely model that three different isomers exist with different lifetimes. On the basis of experimental and theoretical data, a conclusive scheme of the isomerization reaction is presented.

The molecule 3,3(')-diethyl-2,2(')-thiacyanine isomerizes after irradiation with light of the proper wavelength. After excitation, it undergoes a transition, in which one or more conical intersections are involved, back to the ground state to form different product photoisomers. The dynamics before and directly after the transition back to the ground state is investigated by transient absorption spectroscopy in a wavelength region of 360-950 nm, as well as by fluorescence upconversion. It is shown that the excited-state dynamics are governed by two time scales: a short one with a decay time of less than 2 ps and a long one with about 9 ps. A thorough comparison of the experimental results with those of configuration interaction singles and time-dependent density functional theory calculations suggests that these dynamics are related to two competing pathways differing in the molecular twisting on the excited surface after photoexcitation. From the experimental point of view this picture arises taking into account the time scales for ground-state bleach, excited-state absorption, stimulated emission, fluorescence, and assumed hot ground-state absorption both in the solvent methanol and ethylene glycol.

A thorough study of the acid-base behavior of the four histidines and the other titratable residues of the structured domain of human prion protein (125-228) is presented. By using multi-tautomer electrostatic calculations, average titration curves have been built for all titratable residues, using the whole bundles of NMR structures determined at pH 4.5 and 7.0. According to our results, (1) only histidine residues are likely to be involved in the first steps of the pH-driven conformational transition of prion protein; (2) the pK(a)'s of His140 and His177 are approximately 7.0, whereas those of His155 and His187 are < 5.5. 10-ns long molecular dynamics simulations have been performed on five different models, corresponding to the most significant combinations of histidine protonation states. A critical comparison between the available NMR structures and our computational results (1) confirms that His155 and His187 are the residues whose protonation is involved in the conformational rearrangement of huPrP in mildly acidic condition, and (2) shows how their protonation leads to the destructuration of the C-terminal part of HB and to the loss of the last turn of HA that represent the crucial microscopic steps of the rearrangement., ((c) 2006 Wiley-Liss, Inc.)

The main structural and electronic factors playing a role in intramolecular dissociative electron transfer of a simple donor-peptide-acceptor (D-peptide-A) model have been investigated by an integrated computational protocol based on the density functional theory, its time-dependent extension, and the polarizable continuum model. Our results allow us to elucidate the electronic states involved in the process and how they are perturbed by the orientation of the donor and the acceptor with respect to the peptide chain and by the presence of the solvent. We also report a semiquantitative estimation of the rate constant governing electron transfer obtained by a direct quantum mechanical evaluation of all the terms entering the kinetic expressions based on the Marcus theory and its extensions.

We present a computational method, exploiting some features of the polarizable continuum model (PCM) to describe heterogeneous media; it belongs to the family of electrostatic embedding mixed methods, such as the more common quantum-mechanical (QM)/molecular mechanics approaches, with the electrostatic long range effects accounted for by a polarized continuum instead of atomic point charges. Provided effective dielectric constants are determined for the various parts of the system, the method is much faster than its atomistic counterpart, and allows for high-level QM calculations on the fragment of interest, using all the highly efficient computational tools developed for homogeneous PCM. Two case studies (the calculation of the pKa of solvent exposed acidic residues in a model protein, and the calculation of the electron spin resonance spectrum of a typical spin probe partially embedded in a membrane) are analyzed in some detail, to illustrate the application of the method to complex systems.

The excited-state properties of uracil, thymine, and nine other derivatives of uracil have been studied by steady-state and time-resolved spectroscopy. The excited-state lifetimes were measured using femtosecond fluorescence upconversion in the UV. The absorption and emission spectra of five representative compounds have been computed at the TD-DFT level, using the PBE0 exchange-correlation functional for ground- and excited-state geometry optimization and the Polarizable Continuum Model (PCM) to simulate the aqueous solution. The calculated spectra are in good agreement with the experimental ones. Experiments show that the excited-state lifetimes of all the compounds examined are dominated by an ultrafast (<100 fs) component. Only 5-substituted compounds show more complex behavior than uracil, exhibiting longer excited-state lifetimes and biexponential fluorescence decays. The S(0)/S(1) conical intersection, located at CASSCF (8/8) level, is indeed characterized by pyramidalization and out of plane motion of the substituents on the C5 atom. A thorough analysis of the excited-state Potential Energy Surfaces, performed at the PCM/TD-DFT(PBE0) level in aqueous solution, shows that the energy barrier separating the local S(1) minimum from the conical intersection increases going from uracil through thymine to 5-fluorouracil, in agreement with the ordering of the experimental excited-state lifetime.

A general formalism for the calculation of cavitation energies in the framework of the scaled particle theory has been implemented in the Polarizable Continuum Model (PCM), contributing to the nonelectrostatic part of the molecular free energy in solution. The solute cavity and the solvent molecules are described as hard spherocylinders, whose radius and length are related to the actual molecular shape, while the solvent density is estimated from experimental data, or from the solvent molecular volume, suitably scaled. The present model can describe isotropic solutions of spherical and rod-like molecules in spherical or rod-like solvents, and also anisotropic solutions in which the solvent molecules are oriented in space: in this case, the cavitation energy also depends on the relative orientation of solute and solvent molecules. Test calculations have been performed on simple systems to evaluate the accuracy of the present approach, in comparison with other methods and with the available experimental estimates of the cavitation energy, giving encouraging results., ((c) 2005 Wiley Periodicals, Inc.)

The physicochemical effects modulating the conformational behavior and the rate of intramolecular dissociative electron transfer in phthalimide-Aibn-peroxide peptides (n = 0-3) have been studied by an integrated density functional/continuum solvent model. We found that three different orientations of the phthalimide ring are possible, labeled Phihel, PhiC7, and PhipII. In the condensed phase, they are very close in energy when the system is neutral and short. When the peptide chain length increases and the system is negatively charged, Phihel becomes instead the most stable conformer. Our calculations confirm that the 3(10)-helix is the most stable secondary structure for the peptide bridge. However, upon charge injection in the phthalimide end of the phthalimide-Aib3-peroxide, the peptide bridge can adopt an alpha-helix conformation as well. The study of the dependence of the frontier orbitals on the length and on the conformation of the peptide bridge (in agreement with experimental indications) suggests that for n = 3 the process could be influenced by a 3(10) --> alpha-helix conformational transition of the peptide chain.

The role of acidic pH in the conversion of human prion protein to the pathogenic isoform is investigated by means of molecular dynamics simulations, focusing the attention on the effect of protonation of histidine residues on the conformational behavior of human PrPC globular domain. Our simulations reveal a significant loss of alpha-helix content under mildly acidic conditions, due to destructuration of the C-terminal part of HB (thus suggesting a possible involvement of HB into the conformational transition leading to the pathogenic isoform) and a transient lengthening of the native beta-sheet. Protonation of His-187 and His-155 seems to be crucial for the onset of the conformational rearrangement. This finding can be related to the existence of a pathogenic mutation, H187R, which is associated with GSS syndrome. Finally, the relevance of our results for the location of a Cu2+-binding pocket in the C-terminal part of the prion is discussed.

Here we present the first computations of fluorescence spectra in aqueous solution at an accurate quantum mechanical level. From a methodological point of view, our study shows that by only taking into account both bulk effects and explicit solvent molecules it is possible to reproduce solvent effects on the energy and the intensities of the electronic spectra, especially for what concerns pi/pi* transition. The computed absorption and fluorescence spectra are in a good agreement with the available experimental results. The energy ordering between the lowest energy n-pi* and the pi/pi* transitions in uracil strongly depends on the nature of the embedding medium. The geometry of the first solvation shell is remarkably sensitive to the specific electronic state, suggesting that solvent degrees of freedom can act as S1/S2 coupling modes.

The role of intraresidue interactions in determining the conformational behavior of polypeptides is analyzed by means of density functional and post-Hartree-Fock computations on the alanine dipeptide analog and other model compounds. Our computations show that the accuracy of current density functionals is sufficient for H-bond, electrostatic, inductive, and short-range repulsive interactions, whereas medium-range attractions between electron-rich atoms and/or bonds are underestimated. This leads, in turn, to an underestimation of the stability of helical structures w.r.t. extended or folded conformers involving H-bonds. Those results could pave the route for devising local ad hoc corrections able to significantly improve structural and dynamic predictions for polypeptides issuing from DFT computations., (Copyright 2004 Wiley Periodicals, Inc.)

We report a crystallographic and computational analysis of two mutant forms of the Alicyclobacillus acidocaldarius thioredoxin (BacTrx) done in order to evaluate the contribution of two specific amino acids to the thermostability of BacTrx. Our results suggest that the thermostability of BacTrx may be modulated by mutations affecting the overall electrostatic energy of the protein.

The conformational and spectroscopic properties of the tyrosyl radical dipeptide analogue (T(R)DA) are investigated both in gas phase and in aqueous solution by means of density functional calculations. Electronic interactions between backbone and side chain, determining the relative stability of the different energy minimums, depend on the electronic state of the phenoxy substituent. As a consequence, (i) the conformational behavior of T(R)DA is quite different from that of the tyrosine dipeptide analogue, and (ii) the energy required for the homolytic breaking of the OH bond depends on the adopted conformation. The calculated hyperfine coupling constants are in good agreement with the available experimental results. Side-chain-backbone interactions cause an asymmetrization of the magnetic properties of the phenoxy ring and deviations from McConnell relationship. Solvent effects, taken into account by means of a combined discrete/continuum model, significantly affect both the conformational and the magnetic behavior of T(R)DA.

The importance of vicinal and long-range interresidue effects in determining the stability of the collagen triple helix has been investigated by quantum mechanical (QM) and molecular mechanical (MM) computations on suitable model polypeptides, taking into account solvent effects by the polarizable continuum model (PCM). At the QM level, the PII conformation corresponds to an energy minimum for pentapeptide analogues incorporating the sequence Gly-Pro-Pro-Gly, irrespective of the down or up puckering of the pyrrolidine ring. However, our computations indicate that the alternation of down and up prolines characterizing collagen and collagen-like peptides is not due to an intrinsic preference of the Pro-Pro-Gly sequence. This result is confirmed by MM computations of longer polypeptides. Next, MM computations on model triple helices show that a better packing is obtained for specific values of backbone dihedrals, which, in turn, favor the alternation of down and up prolines along each chain.

The conformational behavior of a dipeptide analogue of tyrosine (TDA) has been investigated by density functional methods using the polarizable continuum model (PCM) for the description of solvent effects. Our study points out the interplay of backbone and side chain contributions in determining the relative stabilities of energy minima. In particular, stabilizing interactions between the NH bond and the aromatic ring have a significant effect. The topology of the potential energy surface is significantly modified in aqueous solution due to a general widening of low energy regions and to a stabilization of helical structures.

The conformational behavior of the title compounds has been investigated by Hartree-Fock, MP2, and DFT computations on the most significant structures related to variations of the backbone dihedral angles, cis/trans isomerism around the peptide bond, and diastereoisomeric puckering of the pyrrolidine ring. In vacuum the reversed gamma turn (gammal), characterized by an intramolecular hydrogen bridge, corresponds to the absolute energy minimum for both puckerings (up and down) of the pyrrolidine ring. An additional energy minimum is found in the helix region, but only for an up puckering of the pyrrolidine ring. When solvent effects are included by means of the polarizable continuum model the conformer observed experimentally in condensed phases becomes the absolute minimum. The down puckering is always favored over its up counterpart, albeit by different amounts (0.4-0.5 kcal/mol for helical structures and about 2 kcal/mol for gammal structures). In helical structures cis arrangements of the peptide bond are only slightly less stable than their trans counterparts. This is no longer true for gammal structures, because the formation of an intramolecular hydrogen bond is possible only for trans peptide bonds. In most cases, proline and hydroxyproline show the same general trends; however, the electronegative 4(R) substituent of hydroxyproline leads to a strong preference for up puckerings irrespective of the backbone conformation.

The structure and magnetic properties of one-dimensional chains of representative nitroxides have been studied by a density functional model employing periodic boundary conditions. The optimized geometries are in better agreement with experiments than those obtained from optimizations of model dimeric systems. The spin populations and isotropic hyperfine couplings compare well with the values measured by polarized neutron and electron spin resonance experiments. Magnetic couplings computed by the broken symmetry approach reproduce the ferro- or antiferromagnetic behavior of different nitroxides derived from experiments. These results point out the reliability of the computational model and the significant tuning of all the magnetic properties by intermolecular hydrogen bridges.

Objective: To define the clinical significance of non-organ specific autoantibody positivity in patients in whom routine clinical and laboratory examinations did not detect any disease that might have caused the serological finding., Methods: Out of 1,120 patients consecutively admitted to an outpatient rheumatology clinic, 28 were referred for the evaluation of an autoantibody positivity unrelated to the clinical status. These patients and 28 sex- and age-matched controls underwent a specific work-up with the aim of detecting any underlying infection or autoimmune disease., Results: Eight of the 28 patients (28.5%) were found to be affected by a previously undetected disease: 3 chronic hepatitis C, 3 Sjögren's syndrome, and 2 autoimmune thyroiditis. The remaining 20 did not show any autoimmune or hepatic disease, although 4 of them showed active infection by HBV (n = 1) or HGV (n = 3) and 15 had had a previous infection by hepatotropic viruses (HBV, CMV or EBV). After a follow-up lasting 6-54 months, none of the last 20 patients developed any autoimmune or chronic hepatic disease., Conclusions: A diagnostic work-up is necessary in patients presenting with unexpected autoantibody positivity in order to detect an underlying pre-clinical autoimmune disease and/or unexpected hepatic infection. Patients in whom such a work-up fails to point out any condition should be further followed in order to make an early diagnosis of autoimmune disease.

The importance of local (intraresidue) effects in determining the stability of the collagen triple helix has been investigated with special reference to the role played by hydroxyproline. To this end the dipeptide analogues of L-proline (ProDA), 4(R)-hydroxy-L-proline (HypDA), and 4(R)-fluoro-L-proline (FlpDA) have been studied by means of quantum mechanical ab initio calculations, taking into account solvent effects by the polarizable continuum model (PCM). Our results confirm that the relative stability of up puckerings of the pyrrolidine ring increases with the electronegativity of the 4(R) substituent (X), whereas down puckerings are favored by 4(S) electronegative substituents. Calculations on model compounds show that this effect is due to the interaction between vicinal C-H bonding and C-X antibonding orbitals. Electronegative substituents on the pyrrolidine ring affect cis-trans isomerism around the peptidic bond, with trans isomers stabilized by 4(R) substituents and cis isomers by 4(S) substituents. Also the hydrogen bonding power of the carbonyl moiety following the pyrrolidine ring is affected by 4(R) substituents, but this effect is tuned by the polarity of the embedding medium. Finally, up puckering favors smaller values of the backbone dihedrals phi and psi. All these results strongly support the proposal that the stability of triple helices containing fluorinated or hydroxylated prolines in Y positions is related to the necessity of having up puckerings in those positions.

Fully quantum mechanical calculations exploiting periodic boundary conditions (PBC) have been applied to the study of four different regular structures (alpha- and 3(10)-helix, fully extended and repeated gamma-turns) of the infinite polypeptides of glycine, alanine, and alpha-aminoisobutyric acid (Aib) in vacuo. alpha-Helix is predicted to be the most stable conformer for polyalanine and polyglycine, being stabilized over the 3(10)-helix mainly by more favorable dipole-dipole interactions. Contrary to previous suggestions, steric effects and hydrogen-bond strengths are comparable for both helix structures. 3(10)-Helix is preferred for poly-Aib, since in this case alpha-helix is strongly distorted due to unfavorable intrachain repulsions. Extended structures and repeated gamma-turns are much less stable than helix structures for all of the polypeptides examined, mainly due to the absence of favorable long-range interactions. The optimized geometries are in good agreement with the available experimental data and reveal a remarkable dependence on the nature of the residue forming the polypeptides; at the same time the electronic and structural parameters of each residue strongly depend on the secondary structure of the polypeptides.

The mechanism of long-range electron transfer between the primary and the secondary quinone of photosynthetic reaction centers has been investigated, with particular attention on the role of the iron-histidine bridge. Computations suggest that in such a system, where the molecular subunits are packed together by H-bonds, a mobile electron, injected on one end of the chain, can be carried to the other end by switching the positions of the H-bonded hydrogens. Energy estimates would suggest that the proposed mechanism is plausible and worthy of further experimental investigations., (Copyright 2000 Academic Press.)

Objective: To investigate whether the serological markers of autoimmunity and the clinical features of autoimmune disease which occur in hepatitis C virus (HCV)-infected subjects are correlated to each other and/or to the clinical pattern of the disease., Methods: Seventeen symptom-free, anti-HCV antibody positive subjects, 17 patients with chronic hepatitis C, 21 patients with mixed cryoglobulinemia (MC), and as controls 17 anti-HCV negative patients with dyspepsia were enrolled in a prospective study. A patient history, clinical examination, self-administered questionnaire and laboratory investigations (hepatic enzyme levels, serum HCV-RNA and anti-HCV antibody testing, and serum autoantibody profile) were performed to detect liver and/or autoimmune disease., Results: Serological markers of autoimmunity and clinical findings of autoimmune disease were found to be more frequent in the HCV-infected patients considered as a whole than in controls. However, rheumatoid factor and clinical findings of autoimmune disease were more frequent in MC patients, while anti-smooth muscle antibodies not linked to symptoms or signs of autoimmune disease were detected in all groups of HCV-infected individuals, including healthy carriers and subjects who had recovered from a previous HCV infection., Conclusion: Anti-smooth muscle antibodies, a serological marker of autoimmunity, are detectable in HCV-infected subjects whatever their clinical status. Clinical findings of autoimmune disease prevalently occur in patients with mixed cryoglobulinemia.

The thermophilic bacterium Bacillus thermoantarcticus produces two exocellular polysaccharides (EPS 1 and EPS 2), which can be obtained from the supernatant of liquid cultures by cold-ethanol precipitation, in yields as high as 400 mg liter(sup-1). The EPS fraction was produced with all substrates tested, although a higher yield was obtained with mannose as the carbon and energy source. The EPS content was proportional to the total biomass. On a weight basis, EPS 1 and EPS 2 represented about 27 and 71%, respectively, of the total carbohydrate fraction. EPS 1 is a sulfate heteropolysaccharide containing mannose and glucose in a relative molar proportion of 1.0 and 0.7, respectively. EPS 2 is a sulfate homopolysaccharide containing mannose as the major component. The absolute configurations of hexoses were shown to be d for both EPSs. Nuclear magnetic resonance spectra confirmed the presence of (alpha)-d-mannose and (beta)-d-glucose in EPS 1 and only (alpha)-d-mannose in EPS 2. In addition, (sup1)H nuclear magnetic resonance analysis and chemical analysis indicated the presence of pyruvic acid in EPS 2.

beta-Glycosidases catalyze the synthesis of glycosides when a nucleophilic acceptor other than water is present in the reaction medium. We describe the enzymatyc synthesis of 2-beta-D-galactopyranosyloxyethyl methacrylate (GalEMA) starting from 2-hydroxyethyl methacrylate (HEMA) and p-nitrophenyl-beta-D-galactopyranoside (pNPG) using a beta-glycosidase activity present in the thermophilic archaeon Sulfolobus solfataricus. This thermophilic enzyme catalyzes the transfer of the galactopyranosyl unit from pNPG to the HEMA hydroxyl group by the formation of a new beta-glycosidic bond. The conditions of the biocatalytic system have been optimized to obtain high yield of GalEMA. (c) 1996 John Wiley & Sons, Inc.

Antinuclear antibodies (ANA), as detected by indirect immunofluorescence on HEp-2 cells, have been investigated in five spouses and 41 first-degree relatives of nine probands with polymyositis-dermatomyositis (PM-DM) and in 41 sex- and age-matched controls. ANA were detected in 12 out of the 41 first-degree relatives and in two controls (chi 2 = 6.97; P less than 0.01). HLA typing was done in four out of the nine families; in two of them only, ANA segregated with a haplotype. ANA positivity was not correlated either to sex or to age or to household contact. Our results show that ANA occur in a significant percentage of first-degree relatives of patients with PM-DM. The finding seems to be genetically conditioned.

Among vertebrates, chondrichthyans exhibit peculiar karyotypes and total amount and composition of DNA very different from those of tetrapods and teleosteans. Selachians have relatively large genome sizes (more than 30 pg/N), which are inversely related to the fraction rich in adenine-thymine. Moreover, they show a high chromosome number (2n = 60-100), decreasing in the most specialized species. The karyotypes of Scyliorhinus stellaris, Torpedo ocellata, and T. marmorata have been investigated by several techniques in order to distinguish particular genome fractions along the chromosome arms. C-banding appears to be positive in most telomeric regions in Scyliorhinus and at the interstitial level in the two torpedo species. Studies with restriction enzymes (RE) have just been started in selachians, employing ALU I and HIND III in Scyliorhinus stellaris. The former digests the whole chromosome except the telomeric regions, revealing patterns similar to the C-bands. The latter cuts the chromosomes into several interstitial regions, producing G-bands. Other RE are being studied, which will allow identification in situ of qualitative differences in the various DNAs.