The Purification of Amyloid Fibril Proteins

Amyloid fibril concentrates have been fractionated and shown to have homogeneous fragments of the variable region of immunoglobulin proteins as their major protein constituent. Amyloid fibril protein purification was performed on ten amyloid preparations by sequential gel filtration on Sepharose 4 B and Sephadex G-100 columns equilibrated with 5 M guanidine-HCl in 1 N acetic acid.     

New Neplanocin Analogues. V. A Potent Adenosylhomocysteine Hydrolase Inhibitor Lacking Antiviral Activity. Synthesis And Antiviral Activity Of 6″-Carboxylic Acid Derivatives Of Neplanocin A 1

Abstract

The 6′-carboxylic acid derivative of neplanocin A 3 was synthesized from NPA, and was converted to the corresponding methyl ester 4 and amides 5 and 6. These were evaluated for their anti-RNA-virus activities. Of the derivatives synthesized, only 5 was active against RNA viruses within the concentration range of 0.14-4.88 μg/mL. Compounds 3 and 5 showed a potent inhibitory effect on S-adenosylhomocysteine (AdoHcy) hydrolase from rabbit erythrocytes. Although a close correlation between the inhibitory effect of adenosine analogues on AdoHcy hydrolase and their antiviral potency has been demonstrated, 3 did not show any anti-RNA-virus activities.

     

Contribution of Dysferlin Deficiency to Skeletal Muscle Pathology in Asymptomatic and Severe Dystroglycanopathy Models: Generation of a New Model for Fukuyama Congenital Muscular Dystrophy

Defects in dystroglycan glycosylation are associated with a group of muscular dystrophies, termed dystroglycanopathies, that include Fukuyama congenital muscular dystrophy (FCMD). It is widely believed that abnormal glycosylation of dystroglycan leads to disease-causing membrane fragility. We previously generated knock-in mice carrying a founder retrotransposal insertion in fukutin, the gene responsible for FCMD, but these mice did not develop muscular dystrophy, which hindered exploring therapeutic strategies. We hypothesized that dysferlin functions may contribute to muscle cell viability in the knock-in mice; however, pathological interactions between glycosylation abnormalities and dysferlin defects remain unexplored. To investigate contributions of dysferlin deficiency to the pathology of dystroglycanopathy, we have crossed dysferlin-deficient dysferlin ^sjl/sjl mice to the fukutin-knock-in fukutin ^Hp/− and Large-deficient Large ^myd/myd mice, which are phenotypically distinct models of dystroglycanopathy. The fukutin ^Hp/− mice do not show a dystrophic phenotype; however, (dysferlin ^sjl/sjl: fukutin ^Hp/−) mice showed a deteriorated phenotype compared with (dysferlin ^sjl/sjl: fukutin ^Hp/+) mice. These data indicate that the absence of functional dysferlin in the asymptomatic fukutin ^Hp/− mice triggers disease manifestation and aggravates the dystrophic phenotype. A series of pathological analyses using double mutant mice for Large and dysferlin indicate that the protective effects of dysferlin appear diminished when the dystrophic pathology is severe and also may depend on the amount of dysferlin proteins. Together, our results show that dysferlin exerts protective effects on the fukutin ^Hp/− FCMD mouse model, and the (dysferlin ^sjl/sjl: fukutin ^Hp/−) mice will be useful as a novel model for a recently proposed antisense oligonucleotide therapy for FCMD.     

The primary structure of human gamma-glutamyl transpeptidase

A cDNA hybridizable to that of rat gamma-glutamyl transpeptidase (GGT) was cloned from a cDNA library of human fetal liver. The insert of the cDNA clone contained 1866 bp consisting of an open reading frame (ORF) of 1709 bp (569 amino acids (aa), N-terminal portion truncated) and a 135-bp 3'-untranslated region followed by a polyadenylated tail. In parallel, amino acid sequences of N-terminal portions of heavy and light chains of a purified human GGT were determined. Two stretches of amino acid sequences identical to the N-terminal sequences of heavy and light chains were found in the ORF. We therefore concluded that the clone is a cDNA for human GGT. From the amino acid sequence deduced from cDNA, the heavy and the light chains of the purified enzyme are estimated to be composed of 351 aa (Mr 38,336) and of 189 aa (Mr 20,000), respectively. The heavy chain is preceded by a signal peptide of at least 29 aa presumed to be cleaved by bromelain treatment. Six putative N-glycosylation sites are present in the heavy subunit region and one in the light subunit region. Primary structure and hydrophobicity profile are closely similar to those of rat GGT.     

Genomic Evidence for Speciation with Gene Flow in Broadcast Spawning Marine Invertebrates

How early stages of speciation in free-spawning marine invertebrates proceed is poorly understood. The Western Pacific abalones, Haliotis discus, H. madaka, and H. gigantea, occur in sympatry with shared breeding season and are capable of producing viable F₁ hybrids in spite of being ecologically differentiated. Population genomic analyses revealed that although the three species are genetically distinct, there is evidence for historical and ongoing gene flow among these species. Evidence from demographic modeling suggests that reproductive isolation among the three species started to build in allopatry and has proceeded with gene flow, possibly driven by ecological selection. We identified 27 differentiation islands between the closely related H. discus and H. madaka characterized by high F_ST and d_A, but not high d_XY values, as well as high genetic diversity in one H. madaka population. These genomic signatures suggest differentiation driven by recent ecological divergent selection in presence of gene flow outside of the genomic islands of differentiation. The differentiation islands showed low polymorphism in H. gigantea, and both high F_ST, d_XY, and d_A values between H. discus and H. gigantea, as well as between H. madaka and H. gigantea. Collectively, the Western Pacific abalones appear to occupy the early stages speciation continuum, and the differentiation islands associated with ecological divergence among the abalones do not appear to have acted as barrier loci to gene flow in the younger divergences but appear to do so in older divergences.     

Differences in the Optical Response of MSU and CPP Crystals During Magnetic Orientation: Possibility of Diagnosing Gout and Pseudogout

Pseudogout is crystalline arthritis. It has a similar clinical picture to that of gout, and it is difficult to distinguish the two diseases using conventional analysis methods. However, it is important to identify the different crystals responsible for these two cases because the treatment strategies are different. In a previous study, we reported magnetic orientation of monosodium urate (MSU) crystals, which are the causative agent of gout, at the permanent magnet level. In this study, we investigated the effect of an applied magnetic field on calcium pyrophosphate (CPP) crystals, which are the causative agent of pseudogout, and the difference in the magnetic responses of CPP and MSU crystals. We found that the CPP crystals were oriented in a magnetic field on milli-Tesla order because of the anisotropy of the diamagnetic susceptibility. In addition, the CPP crystals exhibited different anisotropic magnetic properties from those of MSU crystals, which led to a characteristic difference between the orientations of the two crystals. That is, we found that the causative agents of gout and pseudogout responded differently to a magnetic field. This report suggests that the discrimination between CPP and MSU by optical measurements is possible by application of magnetic fields appropriately. © 2023 Bioelectromagnetics Society.     

Histone chaperone activity of Fanconi anemia proteins, FANCD2 and FANCI, is required for DNA crosslink repair

Fanconi anaemia (FA) is a rare hereditary disorder characterized by genomic instability and cancer susceptibility. A key FA protein, FANCD2, is targeted to chromatin with its partner, FANCI, and plays a critical role in DNA crosslink repair. However, the molecular function of chromatin-bound FANCD2-FANCI is still poorly understood. In the present study, we found that FANCD2 possesses nucleosome-assembly activity in vitro. The mobility of histone H3 was reduced in FANCD2-knockdown cells following treatment with an interstrand DNA crosslinker, mitomycin C. Furthermore, cells harbouring FANCD2 mutations that were defective in nucleosome assembly displayed impaired survival upon cisplatin treatment. Although FANCI by itself lacked nucleosome-assembly activity, it significantly stimulated FANCD2-mediated nucleosome assembly. These observations suggest that FANCD2-FANCI may regulate chromatin dynamics during DNA repair.     

Molecular identification of oesophagostomum and Trichuris eggs isolated from wild Japanese macaques

Natural habitat fragmentation and reducing habitat quality have resulted in an increased appearance of Japanese macaques, Macaca fuscata (Gray, 1870), in suburban areas in Japan. To investigate the risk of zoonotic infections, a coprological survey of helminth eggs passed by wild Japanese macaques was carried out in 2009 and 2010 in Shiga Prefecture, Japan. Microscopic examination found helminth eggs in high prevalence, and nucleotide sequencing of DNA extracted from the eggs identified Oesophagostomum cf. aculeatum and Trichuris trichiura. A fecal culture also detected infective larvae of Strongyloides fuelleborni. These zoonotic nematodes pose a potential health issue to local people in areas frequented by Japanese macaques.     

Syntheses of α and γ-BHC Alkylthio Analogs

Meso-(1245/36)-1,2,4,5,6-pentachloro-3-methylthiocyclohexane, and (124/356)-1,2,4,5,6-pentachloro-3-methylthio and ethylthiocyclohexanes were prepared from (1234/56)-1,4,5,6-tetrachloro-2,3-epoxycyclohexane (α-BTC cis-epoxide).