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Temporal criterion for behavior: orientation of the young Japanese flounder Paralichthys olivaceus.

When considering a behavioral pattern as a specific type of mechanism, an inherent problem is that it is difficult to determine to what extent the mechanism is programmed to behave selectively in individual situations. To probe this question further, we investigated the orientation of the body axis of the young Japanese flounder Paralichthys olivaceus. The pattern of the substrate and the orientation of neighboring fish were recognized as the determining factors for orientation in P. olivaceus. It was expected, therefore, that a group of individuals would exhibit a definite orientation pattern with respect to the striped pattern. However, the global orientation patterns on the striped substrate based on ten individuals could be classified into two categories: perpendicular and cross to the stripe pattern. This suggests that the substrate pattern and the surrounding individuals operated as distinct temporal criteria as stimuli for orientation. Analysis based on the local and global viewpoints reveals the temporality quantitatively.

Journal Article↗

Adaptability of endothelial orientation to blood flow dynamics--a morphologic analysis.

Vascular endothelium has a certain specificity in its anatomical orientation. The long axis of the endothelial cell and the shingle effect of cell process overlapping is always oriented parallel to the direction of blood flow. To determine whether this specific anatomical orientation would be maintained when the direction of blood flow was altered, twenty ilio-femoral vein segments and twenty vascular patches of carotid artery and jugular vein were studied. Valvulotomised ilio-femoral vein segments (n = 20) were interposed into adjacent arteries in a non-reversed fashion (the direction blood flow is reversed to the cell process overlapping). One centimeter-square carotid artery and jugular vein patches (n = 20) were rotated 90 degrees (endothelial cell orientation is perpendicular to the direction of blood flow). Reversal of endothelial cell overlapping to complement the complete reversal of the direction of blood flow exists in the vein graft through re-endothelialisation and endothelial reorientation. The process of endothelial reversal occurred in stages (endothelial cell process retraction, interdigitation, complete reversal) and was completed within 8-16 days. In the vascular patch, endothelial re-orientation occurred in 8 days through cell re-orientation and replacement. Mitosis was not encountered in either study. This report highlights certain functions and the adaptive capability of endothelial cells to complement the direction of blood flow.

Adaptation, Physiological↗

Phototactic orientation mechanism in the ciliate Fabrea salina, as inferred from numerical simulations.

The marine ciliate Fabrea salina shows a clear positive phototaxis, but the mechanism by which a single cell is able to detect the direction of light and orient its swimming accordingly is still unknown. A simple model of phototaxis is that of a biased random walk, where the bias due to light can affect one or more of the parameters that characterize a random walk, i.e., the mean speed, the frequency distribution of the angles of directional changes and the frequency of directional changes. Since experimental evidence has shown no effect of light on the mean speed of Fabrea salina, we have excluded models depending on this parameter. We have, therefore, investigated the phototactic orientation of Fabrea salina by computer simulation of two simple models, the first where light affects the frequency distribution of the angles of directional changes (model M1) and the second where the light bias modifies the frequency of directional changes (model M2). Simulated M1 cells directly orient their swimming towards the direction of light, regardless of their current swimming orientation; simulated M2 cells, on the contrary, are unable to actively orient their motion, but remain locked along the light direction once they find it by chance. The simulations show that these two orientation models lead to different macroscopic behaviours of the simulated cell populations. By comparing the results of the simulations with the experimental ones, we have found that the phototactic behaviour of real cells is more similar to that of the M2 model.

Animals↗

Biological and psychosocial determinants of male and female human sexual orientation.

Some propositions on male and female sexual orientation will be considered. Some of these are established; others are more speculative. The aim is to offer some notes towards a coherent, comprehensive theory of sexual orientation. 1. The distinction between butch and femme lesbians seems real rather than a social construct. 2. High levels of prenatal steroid hormones seem to be causally associated with the sexual orientation of butch lesbians. However it is not established whether the causal process operates prenatally or postnatally (or both). This is so because prenatal hormone levels are thought to correlate positively with postnatal hormone levels. And high postnatal hormone levels may facilitate homosexual behaviour as a consequence of sensation-seeking. 3. Male bisexuals also are interpreted to have been exposed to high prenatal testosterone levels. But (for reasons similar to those outlined above in regard to butch lesbians) it is unclear whether these have a direct prenatal effect on the brain or whether they are precursors of high postnatal testosterone levels, which are associated with male bisexual orientation by promoting sensation-seeking behaviour. 4. Postnatal learning processes seem to be causally involved in the sexual orientation of some femme lesbians and some exclusive male homosexuals. 5. Some homosexual men have genes that predispose to their sexual orientation. 6. The same may apply to some lesbians, but such genes have not, as far as I know, been identified. 7. People (of both sexes) who engage in same-sex sexual behaviour may be classified simultaneously in two ways, viz (1) 'active' vs 'passive' and (2) those who do and those who do not engage (or consider engaging) in sex with members of the opposite sex. Ex hypothesi, some of the 'active' ones initiate some of the 'passive' ones. The active ones are driven more by hormones and the passive ones by psychosocial factors. The active males contain a substantial proportion of self-identified bisexuals; and the active females a substantial proportion of self-identified butches. 8. These two active categories (butch lesbians and male bisexuals) share a number of endocrinological, psychological, morphological and behavioural features vis-a-vis their exclusively homosexual and heterosexual peers. Methods of testing some of these ideas are presented.

Androgens↗

Measurement of orientation in organic thin films.

As the potential for applications utilizing oriented thin films grows, so grows the need for accurate, reliable measurements of molecular orientation and surface coverage. Recent work in our laboratory has been directed toward this goal. In this paper the theoretical and experimental effects of surface roughness and the width of the molecular orientation distribution on spectroscopically measured orientation angles are reviewed, the combination of linear and nonlinear spectroscopic techniques for accurate determination of both the mean and width of an orientation distribution is described, and the theory and methodology necessary to obtain orientation-insensitive surface coverage measurements by second harmonic generation for adsorption isotherm and kinetics investigations are reviewed.

Algorithms↗

Structural consequences of ammonia binding to the manganese center of the photosynthetic oxygen-evolving complex: an X-ray absorption spectroscopy study of isotropic and oriented photosystem II particles.

The structure and orientation of the manganese complex in NH3-treated photosystem II (PS II) membrane particles of spinach are being studied by X-ray absorption spectroscopy. On the basis of earlier work by our group, a structure for the tetranuclear manganese complex of PS II, which consists of two di-mu-oxo-bridged binuclear Mn units linked by a mono-mu-oxo group, has been proposed [Yachandra, V. K., et al. (1993) Science 260, 675-679]. The extended X-ray absorption fine structure (EXAFS) of the complex modified by NH3 binding in the S2-state is suggestive of an increase in the Mn-Mn distance of one of these units from 2.72 +/- 0.02 to 2.87 +/- 0.02 A, whereas the Mn-Mn distance of the second unit seems to be unaffected by NH3 treatment. The elongation of one binuclear center could result from the replacement of one bridging mu-oxo by an amido group. The lengthening of one Mn-Mn distance means that, by NH3 treatment, the distance degeneracy of the 2.7 A Mn-Mn EXAFS interaction is removed. Consequently, the orientation of individual binuclear units with respect to the membrane normal becomes resolvable by EXAFS spectroscopy of partially oriented PS II membrane particles. The angle between the normal of the PS II-containing membrane and the Mn-Mn vector is determined to be 67 degrees +/- 3 degrees for the 2.87 A distance and 55 degrees +/- 4 degrees for the 2.72 A distance. Only small effects on position, shape, and orientation dependence of Mn K-edge spectra result from NH3 treatment, indicating that the Mn oxidation state, the symmetry of the Mn ligand environment, and the orientation of the complex remain essentially unaffected in the annealed NH3 S2-state. Therefore, it seems likely that the angles determined for the ammonia-modified manganese complex are similar to the respective angles of the untreated complex. The structure of the manganese complex and its orientation in the membrane are discussed.

Ammonia↗

Orientation of the oxygen-evolving manganese complex in a photosystem II membrane preparation: an X-ray absorption spectroscopy study.

X-ray absorption spectroscopy has been performed on oriented photosystem II membrane particles isolated from spinach. Structural features of the tetranuclear Mn cluster and the orientation of the cluster with respect to the lipid bilayer were determined in both the S1 and S2 states of the Kok cycle. Variation of the sample orientation with respect to the X-ray e-vector yields highly dichroic K-edge and extended X-ray absorption fine structure spectra (EXAFS), indicative of an asymmetric tetranuclear cluster. Mn-Mn vectors at 2.72 and 3.38 A can be resolved from these measurements using quantitative analysis. The 2.72-A vector, consisting of at least two component vectors, is oriented at an average angle of 60 degrees +/- 7 degrees to the membrane normal, with an average of 1.1 +/- 0.1 interactions per Mn atom. The 3.38-A vector, most probably an average of two vectors, makes an angle of 43 degrees +/- 10 degrees with respect to the membrane normal, with an average of 0.45 +/- 0.07 backscatterer per Mn atom. Upon advance to the S2 state, the orientation of these vectors and the average numbers of backscatterers are approximately invariant. Analysis of more subtle features of the EXAFS reveals changes accompanying this S-state advance that are consistent with the oxidation of Mn during this transition. However, the dominant structural features of the oxygen-evolving complex remain constant in the S1 and S2 states. The structure of the Mn complex and the orientation of the complex in the membrane within the context of dichroism of the X-ray absorption data are discussed.

Chloroplasts↗

Orientation of the alpha-helices of apocytochrome c and derived fragments at membrane interfaces, as studied by circular dichroism.

The orientation of the different helical regions of the mitochondrial precursor protein apocytochrome c has been studied using circular dichroism on isolated fragments of this protein associated with oriented films composed of various phospholipids [de Jongh, H. H. J., Goormaghtigh, E., & Killian, J. A. (1994) Biochemistry (preceding article in this issue)]. Both the N and C terminus adopt helical structures in a membrane environment. The middle region can also be helical, but only in the presence of the N-terminal domain of the protein. In the presence of the unsaturated lipids dioleoylphosphatidylcholine and dioleoylphosphatidylglycerol, all three helices are found to have a preferred orientation perpendicular to the membrane normal, whereas in the presence of the saturated lipids dimyristoylphosphatidylcholine and dimyristoylphosphatidylglycerol, the terminal helices are preferentially oriented parallel to the membrane normal. In films composed of dioleoylphosphatidylserine, it is found that the N-terminal helix is oriented preferentially perpendicular, whereas the C-terminal helix is aligned more parallel to the membrane normal. The differences in preferred orientation between the terminal helices are demonstrated by molecular modeling of the helices at a water-lipid interface. The results are discussed in light of the translocation of apocytochrome c over the outer mitochondrial membrane, an important step in the import process of this protein in mitochondria.

Animals↗

Sequence-dependent peptide binding orientation by the molecular chaperone DnaK.

Hsp70-class molecular chaperones interact with diverse polypeptide substrates, but there is limited information on the structures of different Hsp70-peptide complexes. We have used a site-directed fluorescence labeling and quenching strategy to investigate the orientation of different peptides bound to DnaK from Escherichia coli. DnaK was selectively labeled on opposite sides of the substrate-binding domain (SBD) with the fluorescent probe bimane, and the ability of peptides containing N- or C-terminal tryptophan residues to quench bimane fluorescence was measured. Tryptophan-labeled derivatives of the model peptide NRLLLTG bound with the same forward orientation previously observed in the crystal structure of the DnaK(SBD)-NRLLLTG complex. Derivatives of this peptide containing arginine in the C-terminal rather than N-terminal region, NTLLLRG, also bound in the forward direction indicating that charged residues in the flanking regions of the peptide are not the major determinant of peptide binding orientation. We also tested peptides having proline in one (ELPLVKI) or two (ELPPVKI) central positions. Tryptophan derivatives of each of these peptides bound with a strong preference for the reverse direction relative to that observed for the NRLLLTG and NTLLLRG peptides. Computer modeling the peptides NRLLLTG and ELPPVKI in both the forward and reverse orientations into the DnaK(SBD) indicated that differential hydrogen-bonding patterns and steric constraints of the central peptide residues are likely causes for differences in their binding orientations. These findings establish that DnaK is able to bind substrates in both forward and reverse orientations and suggest that the central residues of the peptide are the major determinants of directional preference.

Amino Acid Sequence↗

Orientation and oligomerization specificity of the Bcr coiled-coil oligomerization domain.

The Bcr oligomerization domain, from the Bcr-Abl oncoprotein, is an attractive therapeutic target for treating leukemias because it is required for cellular transformation. The domain homodimerizes via an antiparallel coiled coil with an adjacent short, helical swap domain. Inspection of the coiled-coil sequence does not reveal obvious determinants of helix-orientation specificity, raising the possibility that the antiparallel orientation preference and/or the dimeric oligomerization state are due to interactions of the swap domains. To better understand how structural specificity is encoded in Bcr, coiled-coil constructs containing either an N- or C-terminal cysteine were synthesized without the swap domain. When cross-linked to adopt exclusively parallel or antiparallel orientations, these showed similar circular dichroism spectra. Both constructs formed coiled-coil dimers, but the antiparallel construct was approximately 16 degrees C more stable than the parallel to thermal denaturation. Equilibrium disulfide-exchange studies confirmed that the isolated coiled-coil homodimer shows a very strong preference for the antiparallel orientation. We conclude that the orientation and oligomerization preferences of Bcr are not caused by the presence of the swap domains, but rather are directly encoded in the coiled-coil sequence. We further explored possible determinants of structural specificity by mutating residues in the d position of the coiled-coil core. Some of the mutations caused a change in orientation specificity, and all of the mutations led to the formation of higher-order oligomers. In the absence of the swap domain, these residues play an important role in disfavoring alternate states and are especially important for encoding dimeric oligomerization specificity.

Amino Acid Sequence↗

A buried polar interaction can direct the relative orientation of helices in a coiled coil.

Coiled coils consist of bundles of two or more alpha-helices that are aligned in a parallel or an antiparallel relative orientation. The designed peptides, Acid-p1 and Base-p1, associate in solution to form a parallel, heterodimeric two-stranded coiled coil [O'Shea, E. K., Lumb, K. J., and Kim, P. S. (1993) Curr. Biol. 3, 658]. The buried interface of this complex is formed by hydrophobic Leu residues, with the exception of an Asn residue from each strand that is positioned to engage in a buried polar interaction. Substitution of these buried Asn residues by Leu residues results in a loss of structural uniqueness, as evidenced by a lack of a particular helix orientation in the Acid-Base coiled-coil complex [Lumb, K. J., and Kim, P. S. (1995) Biochemistry 34, 8642]. Here, we alter the positions of the Asn residues in the Acid and Base peptides such that a buried polar interaction is only expected to occur when the helices are in an antiparallel orientation. The resulting peptides, Acid-a1 and Base-a1, associate to form a helical heterodimer, as shown by circular dichroism (CD) and equilibrium sedimentation centrifugation. The helix orientation preference has been measured using covalently linked, disulfide-containing heterodimers in which the constituent peptides are constrained to interact in either a parallel or an antiparallel orientation. Although both the parallel and antiparallel heterodimers form stable, helical structures, the antiparallel heterodimer is the predominant species at equilibrium when the heterodimers are allowed to undergo thiol-disulfide exchange. In addition, the antiparallel heterodimer is more stable to chemical denaturation than the parallel counterpart by approximately 2.3 kcal/mol. These results demonstrate that a single buried polar interaction in the interface between the helices of a coiled coil is sufficient to determine the relative orientation of its constituent helices.

Amino Acid Sequence↗

Molecular orientation of porphyrins accompanying the formation of chain-aligned DNA films.

Porphyrin-containing DNA solid films with several binding orientations were successfully prepared by drying the aqueous solution of porphyrins and DNA in a magnetic field. By the measurement of linear dichroism absorption spectra from three identical spatial directions, the molecular orientations of porphyrins in chain-aligned DNA films were evaluated. Tetra(N-methylpyridinium-4-yl)-porphyrin was found to be bound at the surface of DNA chains like a patch. This orientation is different from the intercalative or groove binding manners observed in solutions. In contrast, tetra(4-sulfonatophenyl)-porphyrin exhibited an orientation perpendicular to the DNA axis, which may be attributed to H-aggregation of porphyrins along the direction parallel to DNA chains. Heme-protein with coiled coil backbone was also aligned along the DNA chains, orienting ferric protoporphyrin parallel to the magnetic field. The major effect for these molecular orientations would be the molecular packing of the rod-disk or the rod-rod systems.

Animals↗

Orientation of iron bleomycin and porphyrin complexes on DNA fibers.

Bleomycin (Blm) is an antitumor agent that requires iron and oxygen for strand cleavage of DNA. In this study, ferric bleomycin, Fe(III)Blm, or the nitric oxide adduct of ferrous bleomycin, ON-Fe(II)Blm, were bound to one-dimensionally oriented DNA fibers. Reductive nitrosylation of Fe(III) complexes took place in situ on B-form DNA fibers. Electron paramagnetic resonance (EPR) spectra were obtained as a function of the angle phi between the magnetic field B and the fiber axis Zf. For comparison, EPR spectra were acquired for ON-Fe(II)TMpyP and ON-Fe(II)TMpyP-Im on oriented DNA fibers, where TMpyP is 5,10,15,20-tetrakis(1-methyl-4-pyridino)porphyrin and Im is imidazole. EPR spectra showed both low-spin Fe(III)Blm and ON-Fe(II)Blm bound to B-form DNA in two slightly different binding orientations in the ratio of 1:0.2. With A-form DNA, a fraction of bound Fe(III)Blm was high spin. Specifically, the angle beta between the fiber axis Zf and the g axis, gz, perpendicular to or nearly perpendicular to the equatorial plane of the iron complex was estimated as 20 degrees and 25 degrees for ON-Fe(II)Blm and 30 degrees and 25 degrees for Fe(III)Blm, respectively. The angle gamma that determines the orientation of gx and gy axes was estimated as 90 degrees for the two ON-Fe(II)Blm species and 10 degrees for the two Fe(III)Blm species, respectively. The NO was held rigidly in place as the temperature increased from 123 K to room temperature for ON-Fe(II)Blm but not for ON-Fe(II)TMpyP or ON-Fe(II)TMpyP-Im. It is hypothesized that the NO is structurally oriented by hydrogen bonding like the peroxide is held in HO2(-)-Co(III)Blm (Wu et al. J. Am. Chem. Soc. 1996, 118, 1281-1294). The EPR parameters are consistent with a six-coordinate complex for ON-Fe(II)Blm, although the superhyperfine structure from the trans nitrogen was not detected. The increase in g value anisotropy upon binding ON-Fe(II)Blm to DNA fiber may be caused by an increase in the overlap of d pi and 2p pi* orbitals induced by an interaction of NO with DNA and/or by a perturbation of d orbitals due to the pyrimidine-guanine interaction. It is concluded that the EPR parameters of ON-Fe(II)Blm and Fe(III)Blm bound to oriented DNA support the hypothesis that FeBlm species bind to DNA with adduct structures similar to those formed by related CoBlm species and DNA.

Animals↗

An analytical solution to the problem of the orientation of rigid particles by planar obstacles. Application to membrane systems and to the calculation of dipolar couplings in protein NMR spectroscopy.

Nonspherical particles or molecules experience an ordering effect in the presence of obstacles due to the restrictions they place on the orientation of those molecules that are in their proximity. Obstacles may be the limits of a membrane in which the molecule is embedded, oriented mesoscopic systems such as bicelles, or membrane fragments used to induce weak protein alignment in a magnetic field. The overall shape of most proteins can be described to a good approximation by an ellipsoidal particle. Here we describe and solve analytically the problem of the orientation of ellipsoidal particles by planar obstacles. Simple expressions are derived for the orientational distribution function and the order parameter. These expressions allow the analytical calculation of the residual dipolar couplings for a protein of known three-dimensional structure oriented by steric effects. The results are in good agreement with experiment and with the results of previously described simulations. However, they are obtained analytically in a fraction of the time and therefore open the possibility to include the optimization of the overall shape in the determination of three-dimensional structures using residual dipolar coupling constraints. The equations derived are general and can also be applied to problems of a completely different nature. In particular, previous equations describing the orientation of particles embedded in membranes are verified and generalized here.

Membrane Proteins↗

Orientation and conformation of cell-penetrating peptide penetratin in phospholipid vesicle membranes determined by polarized-light spectroscopy.

The orientation and conformation of the cell-penetrating peptide "penetratin" associated with phospholipid vesicle membranes has been determined using polarized-light spectroscopy. The magnitude of orientation of penetratin is unprecedented for a solute in our membrane system, which we believe indicates a strong, specific interaction with the membrane. To validate the spectroscopic technique for studying the orientation of the two tryptophan residues of penetratin, we applied tryptophan octyl ester as a model compound. It is found to be incorporated in the membrane and preferentially oriented with its hydrophobic benzene edge of the indole chromophore pointing into the membrane and its hydrophilic groups oriented toward the water. For penetratin, the results indicate that a central alpha-helical part of the peptide is aligned parallel with the membrane surface, while the ends of the peptide adopt a planar structure. The planes of the two tryptophan side chains show a preferred orientation parallel with the membrane surface, indicating that they are not inserted into the membrane.

Carrier Proteins↗

Solid-state NMR and quantum chemical investigations of 13Calpha shielding tensor magnitudes and orientations in peptides: determining phi and psi torsion angles.

We report the experimental determination of the (13)C(alpha) chemical shift tensors of Ala, Leu, Val, Phe, and Met in a number of polycrystalline peptides with known X-ray or de novo solid-state NMR structures. The 700 Hz dipolar coupling between (13)C(alpha) and its directly bonded (14)N permits extraction of both the magnitude and the orientation of the shielding tensor with respect to the C(alpha)-N bond vector. The chemical shift anisotropy (CSA) is recoupled under magic-angle spinning using the SUPER technique (Liu et al., J. Magn. Reson. 2002, 155, 15-28) to yield quasi-static chemical shift powder patterns. The tensor orientation is extracted from the (13)C-(14)N dipolar modulation of the powder line shapes. The magnitudes and orientations of the experimental (13)C(alpha) chemical shift tensors are found to be in good accord with those predicted from quantum chemical calculations. Using these principal values and orientations, supplemented with previously measured tensor orientations from (13)C-(15)N and (13)C-(1)H dipolar experiments, we are able to predict the (phi, psi, chi(1)) angles of Ala and Val within 5.8 degrees of the crystallographic values. This opens up a route to accurate determination of torsion angles in proteins based on shielding tensor magnitude and orientation information using labeled compounds, as well as the structure elucidation of noncrystalline organic compounds using natural abundance (13)C NMR techniques.

Carbon Isotopes↗

Electron paramagnetic resonance study of partially oriented clay platelets intercalated with copper(II) 1,4,8,11-tetraazacyclotetradecane.

Electron paramagnetic resonance (EPR) spectra of powder and oriented films of montmorillonite, hectorite, and saponite intercalated with [Cu(cyclam)](2+) (cyclam = 1,4,8,11-tetraazacyclotetradecane) exhibit three components: an orientation-dependent component without hyperfine features, an orientation-dependent component with hyperfine features, and an orientation-independent component without hyperfine feature. EPR spectra of [Cu(cyclam)](2+)-saponite, which exhibit only two components and the best resolved hyperfine features, were simulated. The spectra indicate that a large portion of the saponite platelets are inclined to the glass surface, although they tend to align with their basal planes parallel to the glass surface. The orientation-dependent spectra could be simulated by introducing a Gaussian distribution with a standard deviation of 20 degrees for the inclination angle. The standard deviation may be used as a disorder parameter for the microcrystals assembled on glass plates. Spectral simulation also shows that the CuN(4) plane of [Cu(cyclam)](2+) is parallel to the clay layers. EPR spectra of some other partially oriented systems are also discussed.

Aluminum Silicates↗

Single-molecule orientations in dyed salt crystals.

We present single-molecule confocal microscopy studies of orientational distributions for luminophores isolated in potassium hydrogen phthalate (KAP) crystals. The incorporation of dye molecules that bear no size or shape similarity to the host ions is observed, demonstrating that single-molecule studies on mixed crystals need not be restricted to isomorphous host/guest pairs. Violamine R is oriented and overgrown by the fast vicinal slopes of growth hillocks within the symmetry-related {010} growth sectors and DCM deposits in the {11} growth sectors of KAP. Both mixed crystals exhibit modest absorption dichroism relative to basic pyranine-doped K(2)SO(4). The latter was studied to ensure that a range of orientations was sampled in our experiments. Average orientations determined at the single-molecule level were in close agreement to ensemble-averaged measurements for all three systems, and the chromophore orientational distributions were broader than anticipated, indicating that the crystals incorporate guest molecules in a range of orientations outside the measured ensemble average.

Journal Article↗