Local mobility within villin 14T probed via heteronuclear relaxation measurements and a reduced spectral density mapping

Villin 14T, a representative domain from the actin severing and bundling protein villin, binds calcium ions and actin monomers. To begin to understand the contributions of mobility to the villin-calcium and villin-actin interactions, relaxation rates for magnetization involving the amide nitrogens and protons have been measured for 15N-labeled villin 14T in solution. Although we have measured the complete set of rates required for a full spectral density map, difficulties in the accurate measurement of relaxation rates for antiphase coherence and two-spin order led us to consider a reduced mapping formalism. From the reduced spectral density map, a model-free analysis, or directly from the measured Nx,y relaxation rates, local variations in mobility along the backbone of villin 14T have been revealed. Fast motions are evident not only at the amino and carboxyl termini but also in the turn between strands beta 4 and beta 5 of the central beta-sheet and in the turn between helix alpha 3 and strand beta 7. Slower motions are suggested for the turn between strands beta 2 and beta 3. Motions on the microsecond to millisecond time scale have been probed directly by examining the dependence of the proton transverse relaxation rate on the spin-locking field strength. Leu11 shows a strong dependence on field strength, implying conformational exchange with a time constant of 125 +/- 69 microseconds. The backbone at the actin-binding interface appears to be rather rigid.     

CaBPs and other immunohistochemical markers of the human vomeronasal system: a comparison with other mammals

After more than two centuries of almost sporadic inquiry as to the existence and function of the human vomeronasal system (VNS), the last decade has seen a resurgent interest in it. The principal question vexing many laboratories is whether adult humans retain the VNS that clearly develops during fetal growth. Additional questions are whether the structurally defined fetal VNS has any function role, and if this structure and function extend into postnatal life. One research tool that has been successfully used to identify key components of the mammalian VNS has been immunohistochemistry (IHC). This technique has clearly defined the vomeronasal receptor neurons in the vomeronasal organ, the vomeronasal nerve that projects into the central nervous system, and the target of this nerve, the accessory olfactory bulb. This review will discuss immunohistochemical studies that have identified these features in the mammalian VNS, and relate them to structural and IHC studies of the fetal and adult human VNS. Suggestions as to future studies to clarify the status of the human VNO also are offered.     

Glucose oxidase from Penicillium amagasakiense. Primary structure and comparison with other glucose-methanol-choline (GMC) oxidoreductases

The complete amino acid sequence of glucose oxidase from Penicillium amagasakiense was determined by Edman degradation and mass spectrometry of peptide fragments derived from three different specific proteolytic digests and a cyanogen bromide cleavage. The complete sequence of each monomer comprises 587 amino acid residues, contains three cysteine residues, and seven potential N-glycosylation sites, of which at least five were confirmed to be glycosylated. Glucose oxidase from P. amagasakiense shows a high degree of identity (66%) and 79% similarity to glucose oxidase from Aspergillus niger, and is a member of the glucose-methanol-choline (GMC) oxidoreductase family. The tertiary structures of glucose oxidase from A. niger and cholesterol oxidase from Brevibacterium sterolicum were superimposed to provide a template for the sequence comparison of members of the GMC family. The general topology of the GMC oxidoreductases is conserved, with the exception of the presence of an active site lid in cholesterol oxidase and the insertion of additional structural elements in the substrate-binding domain of alcohol oxidase. The overall structure can be divided into five distinct sequence regions: FAD-binding domain, extended FAD-binding domain, flavin attachment loop and intermediate region, FAD covering lid, and substrate-binding domain. The FAD-binding and the extended FAD-binding domains are composed of several separate sequence regions. The other three regions each comprise a single contiguous sequence. Four major consensus patterns have been identified, including the nucleotide-binding consensus sequence close to their N-termini. The functions of the two motifs recently selected by the Genetics Computer Group, Madison, Wisconsin, as additional signature patterns of the GMC oxidoreductases are discussed. The other consensus patterns belong to either the FAD-binding or the extended FAD-binding domain. In addition, the roles of conserved residues are discussed wherever possible.     

Refined crystal structure and mutagenesis of human granulocyte-macrophage colony-stimulating factor

We reviewed the clinical and technical outcomes of 25 patients with neuromuscular scoliosis, who were treated by Luque instrumentation and posterior spinal fusion from the upper thoracic spine to L5 between 1981 and 1988. A mean curve correction of 46% was obtained operatively with a mean 8 degrees loss of correction during the follow-up period that ranged from 1.9 to 9.4 years (mean, 5.5). Pelvic obliquity was improved 50% from a mean of 16.1 degrees to a mean of 8.1 degrees in 24 patients for whom data were available. At final follow-up, the mean pelvic obliquity increased to 11.4 degrees with only two patients increasing > 8 degrees. The cause for major postoperative increase in pelvic obliquity was continued anterior spinal growth with torsion of the fusion mass and was not related to changes limited to the L5-S1 motion segment. Posterior fusion and instrumentation from the upper thoracic spine to L5 without anterior fusion provides adequate correction and control of spinal deformity for many patients with cerebral palsy. Those patients with significant growth remaining, or with severe deformities, may benefit by preliminary anterior release and fusion or inclusion of the pelvis and sacrum.     

Binge drinking among college students: a comparison of California with other states

The aim of the present study was to describe the expression and distribution of bone morphogenetic protein (BMP) in odontogenic tumors by immunohistochemistry using monoclonal antibody against bovine BMP (BMPMcAb). Eight types of odontogenic tumors (44 cases), including ameloblastoma (20 cases), cementifying fibroma (8 cases), benign cementoblastoma (5 cases), dentinoma (3 cases), compound odontoma (2 cases), adenomatoid odontogenic tumor (2 cases), calcifying epithelial odontogenic tumor (2 cases) and odontogenic fibroma (2 cases), were studied. The results showed that, according to the immunostaining pattern of BMPMcAb, tumors could be classified into two types: all cementifying fibromas, benign cementoblastomas, dentinomas, odontogenic fibromas, and compound odontomas demonstrated a positive reaction, whereas all ameloblastomas, adenomatoid odontogenic tumors, and calcifying epithelial odontogenic tumors were negative. BMPMcAb-positive odontogenic tumors were those tumors with formation of enamel, dentin, cementum or bone. Therefore, BMP might play an important role in the formation of calcified dental tissues and the development of odontogenic tumors contaning such tissues.     

Chemical structure and thermal properties of initiator tRNA from Euphausia sperba in comparison with those of other eucaryotic initiator tRNAs

The nucleotide sequence of initiator tRNA (tRNAiMet) from Euphausia sperba, which was harvested in the Antarctic Sea, was determined to be pA-G-C-A-G-A-G-U-m1G-m2G-C-G-C-A-G-U-G-G-A-A-G-C-G-U-m2G-C-U-G-G-G-C-C-C-A-U-t6 A-A-C-C-C-A-G-A-G-m7G-U-C-G-G-U-A-G-A-psi-C-G-m1A-A-A-C-U-A-C-U-C-U-C-U-G-C-U-A -C-C-AOH by using post-labeling methods recently developed. The nucleotide sequence was very similar to that of mammalian tRNAiMet except for changes in six bases and three modifications: C16, U55, D47 and m5C48 are replaced by U16, psi 55 and unmodified U47 and C48, respectively. A50-U64 and G52-C62 base pairs of mammalian tRNAiMet are reversed in Euphausia tRNAiMet. In addition, the G49-C65 pair of the former is replaced by a less stable G49-U65 pair in Euphausia tRNAiMet. The sequence homology was compared between Euphausia tRNAiMet and over ten different species of eucaryotic tRNAiMet so far sequenced. The melting temperature of Euphausia tRNAiMet was 72.5 degrees C, which is 4.2 degrees C and 8.3 degrees C lower than those of rat liver and yeast tRNAiMet's, respectively. The origin of the thermal instability of Euphausia tRNAiMet is discussed in comparison of its secondary structure compared with those of other eucaryotic tRNAiMet's.     

Iodination of mouse EGF with chloramine T at 4 degrees C: characterization of the iodinated peptide and comparison with other labelling methods

The BP-1/6C3 molecule expressed by early B lineage cells and some stromal cells has been identified as aminopeptidase A (APA). We have previously demonstrated that IL-7 selectively induces BP-1/APA expression by pre-B cells coincident with their growth. Here we directly demonstrate that BP-1 is preferentially expressed by the proliferating subpopulation of normal B cell progenitors. Furthermore, when non-adherent BALB/c bone marrow cells were incubated with IL-7 in the presence of purified BP-1 antibody, B cell precursor proliferation was markedly inhibited. Modulation of the BP-1/6C3 antigen did not occur in the presence of the BP-1 antibody, but APA enzymatic activity was significantly inhibited. The 6C3 antibody, which recognizes a different epitope on the APA molecule, had no effect on either B cell precursor proliferation or APA enzyme activity. We hypothesized that neutralization of APA by the BP-1 antibody results in inhibition of IL-7 driven B cell precursor proliferation. However, when isolated 14.8+ bone marrow cells were cultured with IL-7 in the presence of the BP-1 antibody, no inhibition of proliferation occurred. This data suggested that the effect of the BP-1 antibody might be related to the action of APA on peptides in the marrow microenvironment which are not present in cultures of isolated B cell precursors. The addition of irradiated non-adherent bone marrow cells to the 14.8+ cell cultures restored the inhibitory effect of the BP-1 antibody. Based on these observations, we propose that APA cleaves a small peptide which serves as a natural inhibitor of B cell precursor proliferation.     

Crystal structure of a B-DNA dodecamer containing inosine, d(CGCIAATTCGCG), at 2.4 A resolution and its comparison with other B-DNA dodecamers

The crystal structure of the dodecamer, d(CGCIAATTCGCG), has been determined at 2.4 A resolution by molecular replacement, and refined to an R-factor of 0.174. The structure is isomorphous with that of the B-DNA dodecamer, d(CGCGAATTCGCG), in space group P2(1)2(1)2(1) with cell dimensions of a = 24.9, b = 40.4, and c = 66.4 A. The initial difference Fourier maps clearly indicated the presence of inosine instead of guanine. The structure was refined with 44 water molecules, and compared to the parent dodecamer. Overall the two structures are very similar, and the I:C forms Watson-Crick base pairs with similar hydrogen bond geometry to the G:C base pairs. The propeller twist angle is low for I4:C21 and relatively high for the I16:C9 base pair (-3.2 degrees compared to -23.0 degrees), and the buckle angles alter, probably due to differences in the contacts with symmetry related molecules in the crystal lattice. The central base pairs of d(CGCIAATTCGCG) show the large propeller twist angles, and the narrow minor groove that characterize A-tract DNA, although I:C base pairs cannot form the major groove bifurcated hydrogen bonds that are possible for A:T base pairs.     

Rest or redistribution thallium myocardial imaging for resting myocardial perfusion: a detailed comparison with regional wall motion

Redistribution thallium-201 (201T1) imaging is the most common method of assessing resting myocardial perfusion. However, the equivalence of a redistribution image and a separate rest injection is unclear. Although the presence of a defect on rest imaging has normally been equated with the presence of a myocardial infarction, it has recently been shown that a significant proportion of fixed defects on exercise-redistribution 201T1 actually represent areas of viable myocardium. This study was a detailed comparison of rest and redistribution imaging in 30 patients undergoing routine exercise 201T1 scanning for the assessment of coronary artery disease. A small dose (15 MBq) of 201T1 was administered at rest following the imaging in three standard planar views. Similar stress images were acquired using a further 50-55 MBq of 201T1 administered at peak effort. Redistribution images were acquired 3-4 h later and equilibrium blood pool ventriculography performed using in vivo labelling with 600 MBq 99Tcm-pertechnetate. Of 150 abnormal segments on the exercise scans, 74 (49%) were identified as being reversible on the redistribution scans and 58 (39%) on the rest images. Only 39 (53%) of these reversible defects were identified on both the redistribution and rest scans. Only 41% of the fixed defects on the redistribution images (32% of fixed defects on the rest images) had abnormal wall motion. Therefore, rest and redistribution images are not equivalent. Both rest and redistribution images significantly overestimate myocardial infarction. This may have significant effects on patient selection for revascularization procedures and therefore all patients having perfusion scintigraphy should also have additional assessment of regional wall motion to allow accurate classification of the functional status of myocardial segments.     

Complications of sedation with midazolam in the intensive care unit and a comparison with other sedative regimens

Small angle X-ray diffraction was used to examine arterial smooth muscle cell (SMC) plasma membranes isolated from control and cholesterol-fed (2%) atherosclerotic rabbits. A microsomal membrane enriched with plasma membrane obtained from animals fed cholesterol for up to 13 weeks showed a progressive elevation in the membrane unesterified (free) cholesterol:phospholipid (C/PL) mole ratio. Beyond 9 weeks of cholesterol feeding, X-ray diffraction patterns demonstrated a lateral immiscible cholesterol domain at 37 degrees C with a unit cell periodicity of 34 A coexisting within the liquid crystalline lipid bilayer. On warming, the immiscible cholesterol domain disappeared, and on cooling it reappeared, indicating that the immiscible cholesterol domain was fully reversible. These effects were reproduced in a model C/PL binary lipid system. In rabbits fed cholesterol for less than 9 weeks, lesser increases in membrane C/PL mole ratio were observed. X-ray diffraction analysis demonstrated an increase in membrane bilayer width that correlated with the C/PL mole ratio. This effect was also reproduced in a C/PL binary lipid system. Taken together, these findings demonstrate that in vivo, feeding of cholesterol causes cholesterol-phospholipid interactions in the membrane bilayer that alter bilayer structure and organization. This interaction results in an increase in bilayer width peaking at a saturating membrane cholesterol concentration, beyond which lateral phase separation occurs resulting in the formation of separate cholesterol bilayer domains. These alterations in structure and organization in SMC plasma membranes may have significance in phenotypic modulation or aortic SMC during early atherogenesis.     

NMR structure of human erythropoietin and a comparison with its receptor bound conformation

The solution structure of human erythropoietin (EPO) has been determined by nuclear magnetic resonance spectroscopy and the overall topology of the protein is revealed as a novel combination of features taken from both the long-chain and short-chain families of hematopoietic growth factors. Using the structure and data from mutagenesis studies we have elucidated the key physiochemical properties defining each of the two receptor binding sites on the EPO protein. A comparison of the NMR structure of the free EPO ligand to the receptor bound form, determined by X-ray crystallography, reveals conformational changes that may accompany receptor binding.     

Crystal structure of a pair of follistatin-like and EF-hand calcium-binding domains in BM-40

BM-40 (also known as SPARC or osteonectin) is an anti-adhesive secreted glycoprotein involved in tissue remodelling. Apart from an acidic N-terminal segment, BM-40 consists of a follistatin-like (FS) domain and an EF-hand calcium-binding (EC) domain. Here we report the crystal structure at 3.1 A resolution of the FS-EC domain pair of human BM-40. The two distinct domains interact through a small interface that involves the EF-hand pair of the EC domain. Residues implicated in cell binding, inhibition of cell spreading and disassembly of focal adhesions cluster on one face of BM-40, opposite the binding epitope for collagens and the N-linked carbohydrate. The elongated FS domain is structurally related to serine protease inhibitors of the Kazal family. Notable differences are an insertion into the inhibitory loop in BM-40 and a protruding N-terminal beta-hairpin with striking similarities to epidermal growth factor. This hairpin is likely to act as a rigid spacer in proteins containing tandemly repeated FS domains, such as follistatin and agrin, and forms the heparin-binding site in follistatin.     

NMR solution structure of the pathogenesis-related protein P14a

The nuclear magnetic resonance (NMR) structure of the 15 kDa pathogenesis-related protein P14a, which displays antifungicidal activity and is induced in tomato leaves as a response to pathogen infection, was determined using 15N/13C doubly labeled and unlabeled protein samples. In all, 2030 conformational constraints were collected as input for the distance geometry program DIANA. After energy-minimization with the program OPAL the 20 best conformers had an average root-mean-square deviation value relative to the mean coordinates of 0.88 A for the backbone atoms N, C(alpha) and C', and 1.30 A for all heavy atoms. P14a contains four alpha-helices (I to IV) comprising residues 4 to 17, 27 to 40, 64 to 72 and 93 to 98, a short 3(10)-helix of residues 73 to 75 directly following helix III, and a mixed, four-stranded beta-sheet with topology +3x, -2x, +1, containing the residues 24-25, 53 to 58, 104 to 111 and 117 to 124. These regular secondary structure elements form a novel, complex alpha + beta topology in which the alpha-helices I, III and IV and the 3(10)-helix are located above the plane defined by the beta-sheet, and the alpha-helix II lies below this plane. The alpha-helices and beta-strands are thus arranged in three stacked layers, which are stabilized by two distinct hydrophobic cores associated with the two layer interfaces, giving rise to an "alpha-beta-alpha sandwich". The three-dimensional structure of P14a provides initial leads for identification of the so far unknown active sites and the mode of action of the protein, which is of direct interest for the generation of transgenic plants with improved host defense properties.     

The efficacy of hysteroscopy: a comparison of women presenting with infertility versus other gynaecological symptoms

The aim of this study was to compare the relative efficacy of hysteroscopy as a management tool in the routine initial assessment of women presenting for investigation of infertility with women presenting for investigation of other general gynaecological symptoms for which it is routinely performed. The results of 400 consecutive completed hysteroscopies performed during the primary investigation of infertility are compared with 400 consecutive completed hysteroscopies undertaken in the investigation of women with other gynaecological symptoms. Abnormalities were detected in 12.3% of the 800 hysteroscopies. Significantly less of the infertility group demonstrated abnormality (8.8%) compared to the general gynaecological group (15.8%) (p = 0.0034). There was no difference in the detection rate between primary and secondary infertility. In patients undergoing the procedure for infertility, the results of the hysteroscopy led to an alteration in management in 5.8% of the entire group and in 65.7% of those in whom an abnormality was detected. In patients undergoing the procedure for general gynaecological symptoms, the results of the hysteroscopy led to an alteration in management in 14.5% of the total group and in 97.2% of those in whom an abnormality was detected (p < 0.0001). Structural abnormality correlated with the presence of histological abnormality in 97.2% of cases. In infertile women, the use of hysteroscopy is supported as part of a comprehensive assessment of female reproductive anatomy.     

A case of glycogen storage disease type Ia with multiple hepatic adenomas and G727T mutation in the glucose-6-phosphatase gene, and a comparison with other mutations previously reported

We report a case of 23-yr-old man with glycogen storage disease (GSD) type Ia complicated by multiple hepatic adenomas. Analysis of the G-6-Pase gene using peripheral blood sample showed this patient to be homozygous for a G-to-T transversion at nucleotide 727 in exon 5. This mutation is prevalent among Japanese patients, suggesting that specific genotypes may correlate with different clinical courses or outcomes.     

Functional diagnosis as a tool in rehabilitation: a comparison of teachers and other employees

Contractile cells in the mammalian lung develop in close association with the outgrowing stem bronchi. Fully differentiated smooth muscle cells are typically found in proximal regions, residing in the substantial muscular walls of the major airways and blood vessels. More distally, cells expressing markers of differentiated smooth muscle cells to a variable degree, and which may therefore possess contractile properties, are to be found scattered around the interstitium. We have investigated the temporal and spatial distribution of smooth muscle lineage markers (smooth muscle myosin mRNA) and of those indicative of contractile function (metavinculin mRNA) in the murine lung. In the smooth muscle layers of the bronchi and major blood vessels, these genes are expressed from the onset of pulmonary budding, concurrently with the appearance of alpha-smooth muscle actin and calponin proteins. During fetal development, smooth muscle-associated genes and proteins are restricted to this committed smooth muscle population. The first signs of myofibroblast or pericyte differentiation become manifest perinatally, when their expression of alpha-smooth muscle actin escalates. In the adult lung, such cells may be readily pin-pointed by their positive reaction for metavinculin mRNA, but, at maturity, they do not always coexpress alpha-smooth muscle actin.     

Effect upon negro digit-symbol performance of anticipated comparison with whites and with other negroes.

Hard and easy digit-symbol tasks were administered to Negro male college students in the South under 3 types of instructions: no test, test with local (i.e., own college) norms, and test with national (i.e., white college) norms. A southern-white-student sample was given the hard task only, with the same 3 types of instructions. Negroes scored highest when told they would be compared with other students at their own college, at an intermediate level when anticipating comparison with national norms, and lowest under nonevaluative instructions. These differences tended to be larger on the hard task than on the easy one. The self-ratings of Negroes indicated strongest concern about performance in the national norms condition. In the white sample, digit-symbol scores were about the same under national-norms and local-norms instructions, and both conditions led to higher scores than did the no test condition. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Operation of the laws of sympathetic magic in disgust and other domains.

Investigated the operation of 2 laws of sympathetic magic in 50 American adults (aged 17–50 yrs) using both measurements in the laboratory and questionnaire response. The 1st law, contagion, holds that "once in contact, always in contact." That is, there can be a permanent transfer of properties from one object (usually animate) to another by brief contact. The 2nd law, similarity, holds that "the image equals the object" and that action taken on an object affects similar objects. It was shown that drinks that had briefly contacted a sterilized, dead cockroach become undesirable and that laundered shirts previously worn by a disliked person were less desirable than those previously worn by a liked or neutral person. The law of similarity was demonstrated by showing that Ss rejected acceptable foods shaped into a form that represents a disgusting object and that Ss were less accurate at throwing darts at pictures of the faces of people they like. Evidence was found for the operation of the laws of sympathetic magic in all Ss studied. The laws of sympathetic magic correspond to the 2 basic laws of association (contiguity and similarity). The parallel is discussed, and a disgust-conditioning study to develop this parallel is reported. (29 ref) (PsycINFO Database Record (c) 2011 APA, all rights reserved)