Functional immobilization of recombinant alkaline phosphatases bearing a glutamyl donor substrate peptide of microbial transglutaminase

Covalent and site-specific protein immobilization catalyzed by microbial transglutaminase (MTG) was investigated using recombinant Escherichia coli alkaline phosphatase (AP) tagged with a glutamyl donor substrate peptide (MLAQGS) of MTG. A polystyrene surface physically coated with beta-casein or bovine serum albumin (BSA) was employed as an MTG-specific surface displaying reactive lysine residues. MTG-mediated protein immobilization through catalytic varepsilon-(gamma-glutamyl)lysine bond formation between the peptide tag of recombinant APs and beta-casein- or BSA-coated surface was verified by the detection of AP activity on the surface. It was found that the length and the insertion position of the peptide tag did not significantly affect the efficacy of enzymatic immobilization of the recombinant APs. On the other hand, pH and ionic strength in the reaction media had crucial effects on the immobilization yields. Interestingly, the optimum pH range of MTG-mediated protein immobilization differed markedly from that for an MTG-catalyzed reaction in aqueous solution. The results suggest that the concentration of reactive species due to electrostatic interaction between the enzyme-substrate intermediate and the protein-adsorbed surface is a key factor governing MTG catalysis at a solid surface.     

Evolved Diversification of a Modular Natural Product Pathway: Apratoxins F and G,Two Cytotoxic Cyclic Depsipeptides from a Palmyra Collection of Lyngbya bouillonii

A collection of Lyngbya bouillonii from Palmyra Atoll in the Central Pacific, a site several thousand kilometers distant from all previous collections of this chemically prolific species of cyanobacterium, was found to contain two new cancer cell cytotoxins of the apratoxin family. The structures of the new compounds, apratoxins F and G, were determined by 1D and 2D NMR techniques in combination with mass spectrometric methods. Stereochemistry was explored by using chromatographic analyses of the hydrolytically released fragments in combination with NMR and optical rotation comparisons with known members of the apratoxin family. Apratoxins F and G add fresh insights into the SAR of this family because they incorporate an N‐methyl alanine residue at a position where all prior apratoxins have possessed a proline unit, yet they retain high potency as cytotoxins to H‐460 cancer cells with IC₅₀ values of 2 and 14 nM , respectively. Additional assays using zone inhibition of cancer cells and clonogenic cells give a comparison of the activities of apratoxin F to apratoxin A. Additionally, the clonogenic studies in combination with maximum tolerated dose (MTD) studies provided insights as to dosing schedules that should be used for in vivo studies, and preliminary in vivo evaluation validated the predicted in vivo efficacy for apratoxin A. These new apratoxins are illustrative of a mechanism (the modification of an NRPS adenylation domain specificity pocket) for evolving a biosynthetic pathway so as to diversify the suite of expressed secondary metabolites.     

Effective highlighting modes of graphical user interfaces in visual information search

Universal Access in the Information Society - This study discusses the preferred highlighting mode for graphical user interface (GUI) design. The study investigated the effects of different...     

Localization and Maintenance of Engrafted Mesenchymal Stem Cells Administered via Renal Artery in Kidneys with Ischemia-Reperfusion Injury

Mesenchymal stem cells (MSCs) are a potential therapeutic tool for preventing the progression of acute kidney injury (AKI) to chronic kidney disease (CKD). Herein, we investigated the localization and maintenance of engrafted human bone marrow-derived MSCs in rats subjected to a renal ischemia-reperfusion injury (IRI) and compared the effectiveness of two intravascular injection routes via the renal artery or inferior vena cava. Renal artery injection of MSCs was more effective than intravenous injection at reducing IRI-induced renal fibrosis. Additionally, MSCs injected through the renal artery persisted in injured kidneys for over 21 days, whereas MSCs injected through the inferior vena cava survived for less than 7 days. This difference may be attributed to the antifibrotic effects of MSCs. Interestingly, MSCs injected through the renal artery were localized primarily in glomeruli until day 3 post-IRI, and they decreased in number thereafter. In contrast, the number of MSCs localized in tubular walls, and the interstitium increased gradually until day 21 post-IRI. This localization change may be related to areas of damage caused by IRI because ischemia-induced AKI leads to tubular cell damage. Taken together, these findings suggest renal artery injection of MSCs may be useful for preventing the progression of AKI to CKD.     

Braille learning materials for Braille reading novices: experimental determination of dot code printing area for a pen-type interface read aloud function

Universal Access in the Information Society - This study clarifies a previously established method in which Braille reading novices obtained Braille information aurally. Therein, users touched the...     

A cell-transverse display algorithm for regular and rectilinear 3D grid data

This paper proposes an efficient direct imaging algorithm for constructing iso-surfaces from regular and rectilinear 3D grid data in scientific and engineering visualization. The basic idea is to generate and draw polygons simultaneously by processing the cells spanned by grids in decreasing order of distance from the current viewpoint. Iso-surfaces are generated in five or six tetrahedrons into which the cells are subdivided, and are sent to a graphics device or drawn into a frame buffer on the fly. The execution order of each of the tetrahedrons is identical and is determined by the current viewpoint. Since the algorithm does not need to store intermediate polyhedral data and does not require a depth buffer memory for hidden surface removal, it is applicable to a large quantity of data on a 3D grid, such as computed tomography (CT) data. It is also particularly powerful for semi-transparent display, because transparency calculation can be reduced to image compositing operations if the polygons are drawn in order of their z-depth from the current viewpoint.