Arginine 206 of the C5a receptor is critical for ligand recognition and receptor activation by C-terminal hexapeptide analogs

C5a is a 74-amino-acid glycoprotein whose receptor is a member of the rhodopsin superfamily. While antagonists have been generated to many of these receptors, similar efforts directed at family members whose natural ligands are proteins have met with little success. The recent development of hexapeptide analogs of C5a has allowed us to begin elucidation of the molecular events that lead to activation by combining a structure/activity study of the ligand with receptor mutagenesis. Removal of the hexapeptide's C-terminal arginine reduces affinity by 100-fold and eliminates the ability of the ligand to activate the receptor. Both the guanidino side chain and the free carboxyl of the arginine participate in the interaction. The guanidino group makes the energy-yielding contact with the receptor, while the free carboxylate negates "electrostatic" interference with Arg-206 of the receptor. It is the apparent movement Arg-206 induced by this set of interactions that is responsible for activation, since conversion of Arg-206 to alanine eliminates the agonist activity of the hexapeptides. Surprisingly, activation is a nearly energy-neutral event and may reflect the binding process rather than the final resting site of the ligand.     

The genome of Treponema pallidum: new light on the agent of syphilis

Covert brain activity related to task-free, spontaneous (i.e. unrequested), emotional evaluation of human face images was analysed in 27-channel averaged event-related potential (ERP) map series recorded from 18 healthy subjects while observing random sequences of face images without further instructions. After recording, subjects self-rated each face image on a scale from "liked" to "disliked". These ratings were used to dichotomize the face images into the affective evaluation categories of "liked" and "disliked" for each subject and the subjects into the affective attitudes of "philanthropists" and "misanthropists" (depending on their mean rating across images). Event-related map series were averaged for "liked" and "disliked" face images and for "philanthropists" and "misanthropists". The spatial configuration (landscape) of the electric field maps was assessed numerically by the electric gravity center, a conservative estimate of the mean location of all intracerebral, active, electric sources. Differences in electric gravity center location indicate activity of different neuronal populations. The electric gravity center locations of all event-related maps were averaged over the entire stimulus-on time (450 ms). The mean electric gravity center for disliked faces was located (significant across subjects) more to the right and somewhat more posterior than for liked faces. Similar differences were found between the mean electric gravity centers of misanthropists (more right and posterior) and philanthropists. Our neurophysiological findings are in line with neuropsychological findings, revealing visual emotional processing to depend on affective evaluation category and affective attitude, and extending the conclusions to a paradigm without directed task.     

Electrophysiological evidence for a postperceptual locus of suppression during the attentional blink

When an observer detects a target in a rapid stream of visual stimuli, there is a brief period of time during which the detection of subsequent targets is impaired. In this study, event-related potentials (ERPs) were recorded from normal adult observers to determine whether this "attentional blink" reflects a suppression of perceptual processes or an impairment in postperceptual processes. No suppression was observed during the attentional blink interval for ERP components corresponding to sensory processing (the P1 and N1 components) or semantic analysis (the N400 component). However, complete suppression was observed for an ERP component that has been hypothesized to reflect the updating of working memory (the P3 component). Results indicate that the attentional blink reflects an impairment in a postperceptual stage of processing.     

Paramagnetic 1H-NMR relaxation probes of stereoselectivity in metalloporphyrin catalyzed olefin epoxidation

Enantioselective catalytic epoxidation of olefins is an important problem from both practical and mechanistic points of view. The origins of chiral induction by asymmetric porphyrin and salen complexes were investigated by FT-NMR T1 relaxation techniques. A new chiral vaulted porphyrin (1) that carries (S)-binaphthyl-L-alanine straps across both faces of the porphyrin macrocycle was synthesized and characterized. (R)-styrene oxide was obtained in > 90% ee in the initial stages of styrene epoxidation with F5PhIO catalyzed by 1-Fe(III)Cl. The transition state for olefin epoxidation with high-valent metal-oxo species was modeled by coordinating epoxides to paramagnetic copper complexes of the corresponding ligands. The epoxide enantiomer that better fit the chiral cavity of the catalyst, as revealed by T1 relaxation measurements, was also the major product of catalytic olefin epoxidation. These results are consistent with the "lock-and-key" mechanism of asymmetric catalysis by metalloporphyrins. The copper complex of a chiral salen ligand showed no differentiation in terms of T1 relaxation rates between the enantiomers of cis-beta-methylstyrene oxide in contrast to the high enantioselectivity observed for catalytic epoxidation.     

Periodic orbits: a new language for neuronal dynamics

A new nonlinear dynamical analysis is applied to complex behavior from neuronal systems. The conceptual foundation of this analysis is the abstraction of observed neuronal activities into a dynamical landscape characterized by a hierarchy of "unstable periodic orbits" (UPOs). UPOs are rigorously identified in data sets representative of three different levels of organization in mammalian brain. An analysis based on UPOs affords a novel alternative method of decoding, predicting, and controlling these neuronal systems.     

Model selection for covariance structures analysis in nursing research

Covariance structures analysis is often used in nursing research to appraise statistical models reflecting complex human health processes. The model selection approach in covariance structures analysis is designed to select the "best" model from a specified set of theoretically defensible, competing alternatives, all of which are viewed as approximations. Model selection criteria explicitly incorporate both model misfit in the population and sampling error to evaluate the set of models. The result is that interpretability of model parameters and goodness-of-fit are enhanced simultaneously. Relative merits of the model selection approach are identified in light of technical concerns, parsimony, and use of scientific theory in nursing.     

Cytotoxicity of advanced glycation endproducts is mediated by oxidative stress

Non-enzymatic glycation of proteins with reducing sugars and subsequent transition metal catalysed oxidations leads to the formation of protein bound "advanced glycation endproducts" (AGEs). They accumulate on long-lived proteins and are for example structural components of the beta-amyloid plaques in Alzheimer's disease. Since the oxidation of glycated proteins as well as the interaction of AGEs with cell surface receptors produces superoxide radicals, it was tested in BHK 21 hamster fibroblast cells and SH-SY5Y human neuroblastoma cells if AGEs can exert cytotoxic effects on cells. Cell viability was assessed with three independent tests: MTT-assay (activity of the mitochondrial respiratory chain), lactate dehydrogenase assay (release of cytoplasmatic enzymes, membrane integrity) and Neutral Red assay (active uptake of a hydrophilic dye). Two model AGEs, chicken egg albumin-AGE and BSA-AGE, both caused significant cell death in a dose-dependent manner. The cytotoxic effects of AGEs could be attenuated by alpha-ketoglutarate and pyruvate, by antioxidants such as thioctic acid and N-acetylcysteine, and by aminoguanidine, an inhibitor of nitric oxide synthase. This suggests that reactive oxygen species as well as reactive nitrogen species contribute to AGE mediated cytotoxicity. Since AGEs accumulate on beta-amyloid plaques in AD over time, they may additionally contribute to oxidative stress, cell damage, functional loss and even neuronal cell death in the Alzheimer's disease brain.     

Psychosexual development of women with congenital adrenal hyperplasia

Women with congenital adrenal hyperplasia (CAH) (N = 31) and their unaffected sisters or female cousins (N = 15) participated in a study of psychosexual development. All participants were > or = 18 years of age (mean age, 25 years; range, 18-40). Comparisons were also made between the CAH women with the salt-wasting (SW) form of the disorder and those with simple virilization (SV). A psychosexual assessment protocol examined six variables: (1) sex assignment at birth (probands only); (2) recalled sex-typed behavior during childhood; (3) gender identity and gender role identification in adulthood; (4) relationship status; (5) sexual orientation in fantasy; and (6) sexual orientation in behavior. Salt-wasting status and sex assignment at birth were also ascertained for the CAH women who either refused to participate in the study (N = 10) or could not be traced (N = 13). Compared to the controls, the women with CAH recalled more cross-gender role behavior and less comfort with their sense of "femininity" during childhood. The two groups did not differ in degree of gender dysphoria in adulthood, although the probands showed more cross-gender role identification. Three of the nonparticipant probands were living, as adults, in the male social role (2 reared from birth as boys and 1 who changed from the female to the male social role during adolescence). The CAH women and the controls did not differ in relationship status (married/cohabiting vs. single). The CAH women had lower rates of exclusive heterosexual fantasy and fewer sexual experiences with men than the controls; however, the CAH women did not have more sexual experiences with women than the controls. Comparisons between the SW and SV revealed several differences: the SW were less likely to be assigned to the female sex at birth, recalled more cross-gender role behavior during childhood, were less likely to be married or cohabiting, and had lower rates of sexual experiences with men. The results were discussed in relation to the effects of prenatal androgens on psychosexual differentiation.     

Cloning of human cDNA encoding a novel heptahelix receptor expressed in Burkitt's lymphoma and widely distributed in brain and peripheral tissues

Using PCR with degenerate primers and screening of a human B-cell lymphoblast cDNA library, a full-length cDNA encoding a 375-amino-acid protein was isolated. It contains seven regions of hydrophobic amino acids probably representing membrane-spanning domains of a novel heptahelix receptor, tentatively named CMKRL2. It shows nearly 30% overall identity with the high-affinity IL8 receptor and similar degree of homology with other chemoattractant receptors, including the "fusin" coreceptors for HIV1. Measurements of various transduction pathways following application of a panel of chemokines to transfected cells failed to evoke any reproducible response. Although the natural ligand for CMKRL2 could, thus, not be identified, receptor expression in spleen and lymph nodes as well as in Burkitt's lymphoma (irrespective of EBV status) supports a functional role in activated B-cells. Receptor message was ubiquitously distributed in normal peripheral tissues and CNS, suggesting that CMKRL2 is expressed in widespread cell populations, such as macrophages and neuroglia.     

Identification of two distinct properties of class II major histocompatibility complex-associated peptides

We have examined the interactions of various peptides with the mouse class II major histocompatibility complex molecule I-Ak. The peptides were derived from the model protein hen egg white lysozyme (HEL). The immunodominant peptide of HEL is a 10-mer, residues 52-61. Our previous work established that this sequence contains the key residues for binding and presentation to T cells. Now we show that the binding of this 10-mer sequence resulted in complexes of I-Ak and peptide that, in SDS/PAGE (without boiling the protein), rapidly dissociated from the component alpha and beta chains. The binding interactions were studied in vitro, by incubating purified I-Ak and radiolabeled peptide, or ex vivo, by using antigen-presenting cells incubated with peptides. Peptides with additional residues at either the amino or carboxyl terminus behaved dramatically differently. Complexes of I-Ak with the longer peptides were stable to SDS/PAGE. Very few amino acid additions result in the change from unstable to stable complexes. The important issue here is that when cultured with HEL, antigen-presenting cells selected the HEL peptides containing the 52-61 sequences that favored stability [Nelson, C. A., Roof, R. W., McCourt, D. W. & Unanue, E. R. (1992) Proc. Natl., Acad. Sci. USA 89, 7380-7383]. Also, from other studies, such sequences correlate with a high immunogenicity of the peptide. We conclude that there are structural features of peptides that change the stability of the class II molecule and that are independent of the "core" peptide seen by the T cells.