Validation of the Russian version of the Hypomania Checklist (HCL-32) for the detection of Bipolar II disorder in patients with a current diagnosis of recurrent depression.

BACKGROUND: There are no validated screening tools for Bipolar Disorder (BD) in Russia. OBJECTIVE: To validate the Russian version of the HCL-32 for the detection of Bipolar II disorder (BD II) in patients with Recurrent Depressive Disorder (RDD). METHODS: 409 patients with a current diagnosis of RDD were recruited. The diagnosis was confirmed by the validated Russian version of the Mini International Neuropsychiatric Interview (MINI). Another investigator interviewed the patients using the НСL-32 questions. RESULTS: The total HCL-32 score in patients with BD II was significantly higher than in patients with RDD: 18.2 (4.22) versus 10.85 (5.81) (p<0.001, d=1447). At the cut-off 14 points the sensitivity was 83.7%, specificity 71.9% (p<0.001). The Cronbach's alpha was 0.887 that means good internal consistency. The best discrimination was achieved with 8 items: decreased need for sleep, less shyness or inhibition, talkativeness, more jokes and puns, jumping thoughts distractibility, exhausting or irritating others and high and more optimistic mood. We proposed the reduced variant of the scale, that includes only these 8 variables, with sensitivity 90.5%, specificity 69.8% (AUC=0.88). CONCLUSIONS: The Russian version of the HCL-32 displayed a good ratio of sensitivity to specificity and can be recommended as a validated screening instrument. An 8-item version of HCL needs further research. LIMITATIONS: Limitations include the specific nature of the sample, the HCL-32 assessment carried out by a psychiatrist, no comparison with other BD screening scales. The results of the 8-item version may be sample and culture dependent.

Tropomodulins and tropomodulin/tropomyosin interactions.

The tropomodulins are a family of proteins that cap the slow-growing (pointed) end of actin filaments and require tropomyosin for optimal function. Tropomodulin is an elongated molecule with a molecular mass of about 40 kDa. The C-terminal half of tropomodulin contains one compact cooperatively melting domain, whereas the N-terminal half has no cooperatively melting structure. The N-terminal half of tropomodulin contains two tropomysin-binding sites and a tropomyosin-dependent actin-binding site, the tropomyosin-independent actin-binding site being located at the C terminus. One tropomodulin molecule binds two tropomyosin molecules, and thus one molecule of tropomodulin is necessary and sufficient for capping at the pointed end. Tropomyosin/tropomodulin interactions are isoform specific. Differences in tropomyosin affinity for the two binding sites in tropomodulin may regulate its correct positioning at the pointed end as well as effectiveness of capping the actin filament.

Capping complex formation at the slow-growing end of the actin filament.

Actin filaments are polar; their barbed (fast-growing) and pointed (slow-growing) ends differ in structure and dynamic properties. The slow-growing end is regulated by tropomodulins, a family of capping proteins that require tropomyosins for optimal function. There are four tropomodulin isoforms; their distributions vary depending on tissue type and change during development. The C-terminal half of tropomodulin contains one compact domain represented by alternating alpha-helices and beta-structures. The tropomyosin-independent actin-capping site is located at the C-terminus. The N-terminal half has no regular structure; however, it contains a tropomyosin-dependent actin-capping site and two tropomyosin-binding sites. One tropomodulin molecule can bind two tropomyosin molecules. Effectiveness of tropomodulin binding to tropomyosin depends on the tropomyosin isoform. Regulation of tropomodulin binding at the pointed end as well as capping effectiveness in the presence of specific tropomyosins may affect formation of local cytoskeleton and dynamics of actin filaments in cells.

The shapes and sizes of two domains of tropomodulin, the P-end-capping protein of actin-tropomyosin.

Tropomodulin, the P-end (slow-growing end)-capping protein of the actin-tropomyosin filament, and its fragment (C20) of the C-terminal half were studied by synchrotron small-angle X-ray scattering, restoring low-resolution shapes using an ab initio shape-determining procedure. Tropomodulin is elongated (115 A long) and consists of two domains, one of 65 A in length and the other being similar to C20 in shape and size if the long axes of the two are tilted by about 40 degrees relative to each other. We propose a model for tropomodulin in association with tropomyosin and actin: the N-terminal half of tropomodulin, a rod, binds to the N-terminus of tropomyosin and the C-terminal triangle domain protrudes from the P-end being slightly bent towards the actin subunit at the end, thereby blocking the P-end.

Folding properties of functional domains of tropomodulin.

Tropomodulin (Tmod) stabilizes the actin-tropomyosin filament by capping the slow-growing end (P-end). The N- and C-terminal halves play distinct roles; the N-terminal half interacts with the N-terminal region of tropomyosin, whereas the C-terminal half interacts with actin. Our previous study (A. Kostyukova, K. Maeda, E. Yamauchi, I. Krieger, and Y. Maéda Y., 2000, Eur. J. Biochem. 267:6470-6475) suggested that the two halves are also structurally distinct from each other. We have now studied the folding properties of the two halves, by circular dichroism spectroscopy and by differential scanning calorimetry of the expressed chicken E-type tropomodulin and its large fragments. The results showed that the C-terminal half represents a single, independently folded unit that melts cooperatively through a two-state transition. In contrast, the N-terminal half lacks a definite tertiary structure in solution. The binding of N11, a fragment that corresponds to the first 91 amino acids of the tropomodulin, to tropomyosin substantially stabilized the tropomyosin. This may indicate that the flexible structure of the N-terminal half of tropomodulin in solution is required for binding to tropomyosin and that the N-terminal half acquires its tertiary structure upon binding to tropomyosin.

Crystallization and preliminary characterization of crystals of the C-terminal half fragment of tropomodulin.

Tropomodulin (40 kDa) stabilizes the actin-tropomyosin filament by capping the P end (slow-growing end). The C-terminal half (C20, 20 kDa), an independently folded domain that is believed to be responsible for the P-end capping, has been crystallized. Crystals grew in the presence of Zn(2+) as the solution pH was increased from 3 towards the pI of the protein. The crystals belong to the trigonal space group R3. They have unit-cell parameters a = b = 69.6, c = 101.3 A (mean values, with a estimated standard deviation of 0.009 A) and diffract to 1.9 A resolution when the frozen crystals were measured at 120 K on a rotating-anode X-ray source at 120 K.

Domain structure of tropomodulin: distinct properties of the N-terminal and C-terminal halves.

The structure of tropomodulin, the unique capping protein for the pointed end (the slow-growing end) of an actin filament, was studied. An improved Escherichia coli expression system for chicken E-tropomodulin was established and tropomodulin was prepared, Tmod (N39), in which 15 amino acid residues from the original C-terminus are deleted at the DNA level. This expression and purification system accidentally co-produces an 11-kDa fragment with the original N-terminus (N11). By applying limited proteolysis to Tmod (N39), a 20-kDa C-terminal fragment (C20) was obtained. The limited proteolysis data, as well as the fluorescence spectrometry and CD analyses of Tmod (N39), C20 and N11, revealed that tropomodulin is an alpha-helical protein that consists of two distinct domains. The C-terminal half (20 kDa) is resistant to proteolysis, which suggests that this domain is tightly folded. In contrast, the N-terminal half is susceptible to proteolysis, indicating that in solution this half is likely to be extended or to form a highly flexible structure. Cross-linking experiments with glutaraldehyde indicated that Tmod (N39) and N11 can form complexes with tropomyosin, whereas C20 cannot. This confirms the previous report that the site(s) of interaction with tropomyosin resides in the N-terminal 11-kDa region of tropomodulin.

Investigation of structure and antigenic capacities of Thermococcales cell envelopes and reclassification of "Caldococcus litoralis" Z-1301 as Thermococcus litoralis Z-1301.

Fourteen strains of hyperthermophilic organotrophic anaerobic marine Archaea were isolated from shallow water and deep-sea hot vents, and four of them were characterized. These isolates, eight previously published strains, and six type strains of species of the order Thermococcales were selected for the study of cell wall components by means of thin sectioning or freeze-etching electron microscopy. The cell envelopes of most isolates were shown to consist of regularly arrayed surface protein layers, either single or double, with hexagonal lattice (p6) symmetry, as the exclusive constituents outside the cytoplasmic membrane. The S-layers studied differed in center-to-center spacing and molecular mass of the constituent protein subunits. Polyclonal antisera raised against the cells of 10 species were found to be species-specific and allowed 12 new isolates from shallow water hot vents to be identified as representatives of the species Thermococcus litoralis, Thermococcus stetteri, Thermococcus chitonophagus, and Thermococcus pacificus. Of the 7 deep-sea isolates, only 1 was identified as a T. litoralis strain. Thus, hyperthermophilic marine organotrophic isolates obtained from deep-sea hot vents showed greater diversity with regard to their S-layer proteins than shallow water isolates.

Nucleoside diphosphate kinase from haloalkaliphilic archaeon Natronobacterium magadii: purification and characterization.

An ATP-binding protein from the haloalkaliphilic archaeon Natronobacterium magadii was purified and characterized by affinity chromatography on ATP-agarose and by fast protein liquid chromatography (FPLC) on a Mono Q column. The N-terminal 20 amino acid sequence of the kinase showed a strong sequence similarity of this protein with nucleoside diphosphate (NDP) kinases from different organisms and, accordingly, we believe that this protein is a nucleoside diphosphate kinase, an enzyme whose main function is to exchange gamma-phosphates between nucleoside triphosphates and diphosphates. Comparison of the molecular weights of the NDP kinase monomer determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) (23,000) and of the oligomer determined by sedimentation equilibrium experiments (125,000) indicated that the oligomer is a hexamer. The enzyme was autophosphorylated in the presence of [gamma-32P]ATP, and Mg2+ was required for the incorporation of phosphate. The kinase preserved the ability to transfer gamma-phosphate from ATP to GDP in the range of NaCl concentration from 90 mM to 3.5 M and in the range of pH from 5 to 12. It was found and confirmed by Western blotting that this kinase is one of the proteins that bind specifically to natronobacterial flagellins. NDP kinase from haloalkaliphiles appeared to be simple to purify and to be a suitable enzyme for studies of structure and stability compared with NDP kinases from mesophilic organisms.

Thermococcus gorgonarius sp. nov. and Thermococcus pacificus sp. nov.: heterotrophic extremely thermophilic archaea from New Zealand submarine hot vents.

Two extremely thermophilic archaea, designated W-12 and P-4, were isolated from a geothermal vent in the tidal zone of Whale Island, New Zealand, and from geothermally heated bottom deposits of the Bay of Plenty, New Zealand, respectively. Cells of isolate W-12 are irregular cocci, 0.3-1.2 microns in diameter, motile with polar flagella. The cell envelope consists of one layer of subunits with a major protein of M(r) 75,000. Cells produce protrusions of different kinds: prostheca-like, chains of bubbles, or network of fimbriae. Cells of isolate P-4 are regular cocci, 0.7-1.0 micron in diameter, motile with polar flagella. The cell envelope consists of two layers of subunits; its major protein has an M(r) of 56,000. Both organisms are obligate anaerobes, fermenting peptides in the case of strain W-12, or peptides and starch in the case of P-4. Elemental sulfur is required for growth and is reduced to hydrogen sulfide. The optimal growth temperature of the new isolates is in the range 80-88 degrees C. The optimal growth pH is 6.5-7.2. The G + C content of the DNA of strain W-12 is 50.6 mol%, and of strain P-4 is 53.3 mol%. Based on physiological characteristics, 165 rDNA sequence comparison and DNA base composition, the new isolates were considered to be members of the genus Thermococcus. The low level of DNA-DNA hybridization with the type strains of other Thermococcus species confirms the novel species status of the new isolates. The new isolates are described as Thermococcus gorgonarius sp. nov., with type strain W-12 (= DSM 10395T), and Thermococcus pacificus sp. nov., with type strain P-4 (= DSM 10394T).

Isolation and characterization of flagella and flagellin proteins from the Thermoacidophilic archaea Thermoplasma volcanium and Sulfolobus shibatae.

Isolated flagellar filaments of Sulfolobus shibatae were 15 nm in diameter, and they were composed of two major flagellins which have M(r)s of 31,000 and 33,000 and which stained positively for glycoprotein. The flagellar filaments of Thermoplasma volcanium were 12 nm in diameter and were composed of one major flagellin which has an M(r) of 41,000 and which also stained positively for glycoprotein. N-terminal amino acid sequencing indicated that 18 of the N-terminal 20 amino acid positions of the 41-kDa flagellin of T. volcanium were identical to those of the Methanococcus voltae 31-kDa flagellin. Both flagellins of S. shibatae had identical amino acid sequences for at least 23 of the N-terminal positions. This sequence was least similar to any of the available archaeal flagellin sequences, consistent with the phylogenetic distance of S. shibatae from the other archaea studied.

Unfolding of tertiary structure of Halobacterium halobium flagellins does not result in flagella destruction.

The structure of Halobacterium halobium R1M1 flagella is investigated by the methods of scanning microcalorimetry, circular dichroism, and electron microscopy. It is shown that melting curves of flagella in solutions with a different concentration of NaCl display only one peak of heat capacity that corresponds to one cooperatively melting domain. It is found that flagella do not dissociate after melting. The possible structural organization of archaebacterial flagella is discussed.

Architectonics of a bacterial flagellin filament subunit.

Flagellins of two Escherichia coli strains and their tryptic fragments were studied by different methods. Probabilities of secondary structure formation were also calculated for all flagellins with a known primary structure. The obtained data permit one to suggest a model for the flagellin molecule consisting of a central part responsible for antigenic properties and terminals responsible for polymerization. The central part is variable in length from a few amino acid residues to three-four hundred depending on the bacterial species. The terminal parts consist of about 160 amino acid residues from the N-end and 100 from the C-end.

Flagellin parts acquiring a regular structure during polymerization are disposed on the molecule ends.

Flagellins of two Escherichia coli strains have been investigated by limited proteolysis and scanning microcalorimetry. It has been shown that a monomer flagellin consists of two parts: a central one cooperatively melting, rather resistant to proteases, and the other without a stable tertiary structure and thus easily degrading terminals. Just these terminals acquire a regular structure during polymerization. Core fragments of the two strains have been isolated and characterized.

Domain structure of flagellin.

The chemotaxis of bacteria such as Salmonella and Escherichia coli involves smooth swimming punctuated by periods of tumbling. In smooth swimming the flagellar filaments are left-handed, in tumbling they are right-handed with a different wavelength. The filaments are constructed from a globular protein, flagellin, by a process of self-assembly. The existing models assume that the flagellin molecule is bistable and longitudinal rows of subunits take one of the two possible conformations. Such a model explains the observed different morphology of the flagellum. We have studied Salmonella and E. coli flagellins in polymeric and monomeric forms by scanning microcalorimetry and circular dichroism. We have inferred that a flagellin molecule consists of several domains, two of which are structured at physiological temperatures and are in the monomeric form, while the others acquire a regular form only in the process of polymerization. This phenomenon may be the basis of a process during which the flagellin molecule, fitting into the flagellum, acquires a conformation analogous to that of the neighbouring molecule in the longitudinal row.