Polyaddition of bifunctional spiro orthoesters with bifunctional acid chlorides accompanying double ring‐opening isomerization

Polyaddition of bifunctional spiro orthoesters (SOEs) with bifunctional acid chlorides was examined to develop zero‐shrinkage polymerization. The polyaddition afforded the corresponding polyether‐esters by repeating the addition reaction accompanying the double ring‐opening isomerization of the SOE moiety in a manner similar to the reaction of monofunctional SOEs with acid chlorides. The polyaddition accompanied a slight shrinkage or expansion in volume. © 2000 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 38: 68–73, 2000     

Functional Analysis of the GPI Transamidase Complex by Screening for Amino Acid Mutations in Each Subunit

Glycosylphosphatidylinositol (GPI) anchor modification is a posttranslational modification of proteins that has been conserved in eukaryotes. The biosynthesis and transfer of GPI to proteins are carried out in the endoplasmic reticulum. Attachment of GPI to proteins is mediated by the GPI-transamidase (GPI-TA) complex, which recognizes and cleaves the C-terminal GPI attachment signal of precursor proteins. Then, GPI is transferred to the newly exposed C-terminus of the proteins. GPI-TA consists of five subunits: PIGK, GPAA1, PIGT, PIGS, and PIGU, and the absence of any subunit leads to the loss of activity. Here, we analyzed functionally important residues of the five subunits of GPI-TA by comparing conserved sequences among homologous proteins. In addition, we optimized the purification method for analyzing the structure of GPI-TA. Using purified GPI-TA, preliminary single particle images were obtained. Our results provide guidance for the structural and functional analysis of GPI-TA.     

Recognition of Amphiphiles with Many Pendent Galactose Residues byRicinus communisAgglutinin

Amphiphiles which carry many pendent galactose residues as side chains were prepared by telomerization of 2-methacryloyloxyethyl β-D-galactopyranoside (MEGal) or 3-(2-methacryloyl aminoethylthio)propylD-galactopyranoside (MEPGal, α:β = 3.9:1) using a lipophilic radical initiator. The galactose-carrying amphiphiles (DP (degree of polymerization) = 15) incorporated in liposomes were recognized by a lectin fromRicinus communis(RCA₁₂₀), which was proven by the increase in turbidity of the liposome suspension after mixing with the lectin. The recognition was largely affected by the distance between the galactose residues and the polymer main chain, and the surface density of the amphiphile in the liposomes. The liposomes containing these galactolipids were not taken up by mouse peritoneal macrophages, probably due to a steric hindrance of polymer main chains from the uptake of D-galactose receptors on the macrophages.     

Inhibition of Influenza Virus Replication in MDCK Cells by Circular Dumbbell RNA/DNA Chimeras with Closed Alkyl Loop Structures

We have demonstrated that a new type of circular dumbbell RNA/DNA chimeric oligonucleotide (CDRDON) with two closed nucleotide or alkyl loop structures (hexa‐ethylene glycol) inhibits influenza virus A replication in MDCK cells. The enzymatic synthesis of circular dumbbell RNA/DNA chimeric oligonucleotides was achieved by enzymatically ligating a self‐complementary phosphorylated oligonucleotide with T4‐RNA ligase. The CDRDON‐Al, with two closed alkyl loop structures, showed higher nuclease resistance, hybridization, and cellular uptake than the anti‐S‐ODN and the CDRDON, with two closed nucleotide hairpin‐loop structures. The circular dumbbell RNA/DNA chimeric oligonucleotide (CDRDON‐Al‐PB2‐as), containing an AUG initiation‐codon sequence as the target of PB2, showed highly inhibitory effects on influenza A virus RNA expression. The limited toxicity of unmodified phosphodiester oligonucleotides and the sequence‐specific binding to target mRNA indicate that circular dumbbell RNA/DNA chimeric phosphodiester oligonucleotides can be used with intact cells, and may prevent viral replication in culture.