Starmaker exhibits properties of an intrinsically disordered protein

Fish otoliths composed of calcium carbonate and an organic matrix play a primary role in gravity sensing and the perception of sound. Starmaker (Stm) was the first protein found to be capable of influencing the process of biomineralization of otoliths. Stm dictates the shape, size, and selection of calcium carbonate polymorphs in a concentration-dependent manner. To facilitate exploration of the molecular basis of Stm function, we have developed and optimized a protocol for efficient expression and purification of the homogeneous nontagged Stm. The homogeneous nontagged Stm corresponds to its functional form, which is devoid of a signal peptide. A comprehensive biochemical and biophysical analysis of recombinant Stm, along with in silico examinations, indicate for the first time that Stm exhibits the properties of intrinsically disordered proteins. The functional significance of Stm having intrinsically disordered protein properties and its possible role in controlling the formation of otoliths is discussed.     

Preparation of poly(ethylene glycol) protected nanoparticles with variable bioconjugate ligand density

Maleimide-functional poly(ethylene glycol)-b-poly(epsilon-caprolactone) nanoparticles (NPs) were prepared via the Flash NanoPrecipitation technique. Subsequent reaction with a model ligand, bovine serum albumin (BSA), was conducted using thiol-maleimide conjugation. Reaction of up to 22% of NP surface maleimide-PEG tethers was obtained, with the percent conversion being essentially independent of the ratio of maleimide-PEG to methyl-PEG over the range 30-100%, respectively. At the highest surface coverage, BSA is calculated to essentially cover the NP surface area. Reaction parameters (reaction order and docking constant) describing the extent of ligand conjugation were determined. The reaction order is applicable to the conjugation of ligands presenting free thiol functionalities, while the value of the docking constant is ligand-dependent and accounts for physical and dynamic properties of the ligand-PEG interaction. Jointly, the particle formation process, using block copolymer-directed kinetically controlled assembly and surface functionalization represent a versatile new platform for the preparation of bioconjugated NPs with accurate control of ligand density and minimal processing steps.     

Development of a Culturally Relevant Body Image Instrument among Urban African Americans

Objective: To validate a culturally relevant body image instrument among urban African Americans through three distinct studies. Research Methods and Procedures: In Study 1, 38 medical practitioners performed content validity tests on the instrument. In Study 2, three research staff rated the body image of 283 African‐American public housing residents (75% women, mean age = 44 years), with the residents completing body image, BMI, and percentage body fat measures. In Study 3, 35 African Americans (57% men, mean age = 42) completed body image measures and evaluated their cultural relevance. Results: In Study 1, 97% to 100% of practitioners sorted the jumbled figures into the correct ascending order. The correlation between the body image figures and the practitioners’ weight classifications of the figures was high (r = 0.91). In Study 2, observers arrived at similar ratings of body size with excellent consistency (α = 0.95). Ratings of body image were strongly correlated with participant BMI (r = 0.89 to 0.93 across observers and 0.81 for all participants) and percentage of body fat (r = 0.77 to 0.89 across observers and 0.76 for all participants). In Study 3, body image ratings with the new scale were positively correlated with other validated figural scales. The majority of participants reported that figures in the new body image scale looked most like themselves and other African Americans and were easiest to identify themselves with. Discussion: The instrument displayed strong psychometric performance and cultural relevance, suggesting that the scale is a promising tool for examining body image and obesity among African Americans.     

Chemical and pharmacological investigations of Metaxya rostrata

In a bioassay-guided approach the chemical composition of rhizomes of Metaxya rostrata (Kunth C. Presl) was studied for the first time. Investigations of the cytotoxicity of extracts and fractions on SW480 colorectal carcinoma cells resulted in the isolation of two polyphenols--cinnamtannin B-1 and aesculitannin B. The structures of the compounds were elucidated by different NMR experiments. Additionally, sugars, common sterols, such as sitosterol, stigmasterol and campesterol, as well as chlorogenic acid and caffeic acid were identified in Metaxya rostrata.     

High-throughput identification of fucosyltransferase inhibitors using carbohydrate microarrays

A noncovalent carbohydrate microarray was used to screen for possible inhibitors to fucosyltransferases, which are critical to the synthesis of inflammation mediators like sialyl Lewis x (SLe(x)). Inhibition was followed by observation of the transferred fucose on the carbohydrate array with the lectin Tetragonolobus purpureas. Of these compounds, four inhibitors with nanomolar Ki(s) were discovered, with three of the top five inhibitors exhibiting a common architecture.     

A model of the acid sphingomyelinase phosphoesterase domain based on its remote structural homolog purple acid phosphatase

Sequence profile and fold recognition methods identified mammalian purple acid phosphatase (PAP), a member of a dimetal-containing phosphoesterase (DMP) family, as a remote homolog of human acid sphingomyelinase (ASM). A model of the phosphoesterase domain of ASM was built based on its predicted secondary structure and the metal-coordinating residues of PAP. Due to the low sequence identity between ASM and PAP (approximately 15%), the highest degree of confidence in the model resides in the metal-binding motifs. The ASM model predicts residues Asp 206, Asp 278, Asn 318, His 425, and His 457 to be dimetal coordinating. A putative orientation for the phosphorylcholine head group of the ASM substrate, sphingomyelin (SM), was made based on the predicted catalysis of the phosphorus-oxygen bond in the active site of ASM and on a structural comparison of the PAP-phosphate complex to the C-reactive protein-phosphorylcholine complex. These complexes revealed similar spatial interactions between the metal-coordinating residues, the metals, and the phosphate groups, suggesting a putative orientation for the head group in ASM consistent with the mechanism considerations. A conserved sequence motif in ASM, NX3CX3N, was identified (Asn 381 to Asn 389) and is predicted to interact with the choline amine moiety in SM. The resulting ASM model suggests that the enzyme uses an SN2-type catalytic mechanism to hydrolyze SM, similar to other DMPs. His 319 in ASM is predicted to protonate the ceramide-leaving group in the catalysis of SM. The putative functional roles of several ASM Niemann-Pick missense mutations, located in the predicted phosphoesterase domain, are discussed in context to the model.     

Emerging roles for phospholipid transfer protein in lipid and lipoprotein metabolism

PURPOSE OF REVIEW: This review highlights the recent key advances in our understanding of the role of phospholipid transfer protein in lipid and lipoprotein metabolism. RECENT FINDINGS: The overexpression of human phospholipid transfer protein in mice is associated with an increase in atherosclerosis. This is consistent with earlier studies using mouse models suggesting that phospholipid transfer protein was pro-atherogenic. The presence of phospholipid transfer protein in macrophages and atherosclerotic lesions suggests that it could be either anti-atherogenic by facilitating lipid efflux or pro-atherogenic by facilitating lipid retention. Phospholipid transfer protein may also be a key player in reverse cholesterol transport, as it interacts with the adenosine triphosphate-binding cassette transporter A1 and facilitates lipid efflux from peripheral cells. Both the release of chymase, a neutral protease, from mast cells and the oxidation of HDL by hypochlorous acid can impair the function of phospholipid transfer protein in reverse cholesterol transport. Studies of phospholipid transfer protein-mediated phospholipid transfer activity in humans support a role for phospholipid transfer protein in hypertriglyceridemia, obesity, diabetes, inflammation and coronary artery disease, and in the modulation of LDL particle density and size. Furthermore, recent evidence suggests that phospholipid transfer protein may play a role in reproductive processes, in lipid and lipoprotein metabolism in the central nervous system, and in neurodegenerative disease. SUMMARY: Phospholipid transfer protein is emerging as a multifaceted and multifunctional player in lipid and lipoprotein metabolism, but much additional work will be required to understand the significance of these recent findings for clinical practice.     

A care perspective on coercion and autonomy

In the Netherlands there is a growing debate over the possibility of introducting 'compassionate interference' as a form of good psychiatric care. Instead of respecting the autonomy of the patient by adopting an attitude of non-interference, professional carers should take a more active and commited role. There was a great deal of hostile reaction to this suggestion, the most commonly voiced criticism being that it smacked of 'modern paternalism'. Still, the current conception of care leaves us with a paradox. On the one hand patients are regarded as individuals who have a strong interest in ( and a right to) freedom and non-interference; on the other hand many of them have a desperate need for flourishing, viable relationships. In fact, part of their problem is that they cannot relate very well with other people. This creates a dichotomy, because respecting patients' autonomy often means that they cannot be given the help they so desperately need. In this respect current care practices do not answer the caring needs of these patients.The criticism on care practices is to be considerd as important. It invites us to reexamine and reevaluate the current conception of caring relationships and its main values. In line with this reexamination an alternative perspective on care is introduced in this paper, a perspective in which `compassionate interference' is not so much a threat to autonomy, but a means of attaining autonomy. For this we need a different definition of autonomy than that commonly used in current care practice.     

The recombinant limb as a model for the study of limb patterning, and its application to muscle development

The recombinant limb is a model system that has proved fruitful for analyzing epithelial-mesenchymal interactions and understanding the functional properties of the components of the limb bud. Here we present an overview of some of the insights obtained through the use of this technique. Among these are the understanding that fore or hind limb identity is inherent to the limb bud mesoderm, that the apical ectodermal ridge (AER) is a permissive signaling center and that the limb bud ectoderm plays a central role in the control of dorsoventral polarity. Recombinant limb studies have also allowed the identification of the affected tissue component in several limb mutants. More recently this model has been applied to the study of regulation of gene expressions related to patterning. In this report we use recombinant limbs to analyze pattering of the Pax3 expressing limb muscle cell lineage in the early stages of limb development. In recombinant limbs made without the zone of polarizing activity (ZPA), myoblasts appear intermingled with other mesodermal cells at the beginning of the recombinant limb development. Rapidly thereafter, the muscle precursors segregate and organize around the central forming chondrogenic core of the recombinant. Although this segregation is reminiscent of that occurring during normal development, the myoblasts in the recombinant fail to proliferate appropriately and also fail to migrate distally. Consequently, the muscle pattern in the recombinant limb is defective indicating that normal patterning cues are absent. However, recombinant limbs polarized with a ZPA exhibited a larger mass of muscle cells and a more normal morphogenesis, supporting a role for this signaling center in limb muscle development. Finally, we have ruled out host somite contributions to recombinant limbs by grafting chick recombinant limbs to quail hosts. This initial report demonstrates the value of the recombinant limb model system for dissecting the environmental cues required for normal muscle limb patterning. Received: 31 August 1998 / Accepted: 29 September 1998