Molecular and catalytic properties of 2,4'-dihydroxyacetophenone dioxygenase from Burkholderia sp. AZ11

The gene dad encoding 2,4'-dihydroxyacetophenone (DHAP) dioxygenase was cloned from Burkholderia sp. AZ11. The initiation codon GTG was converted to ATG for high-level expression of the enzyme in Escherichia coli. The enzyme was moderately thermostable, and the recombinant enzyme was briefly purified. The enzyme (M(r)=90 kDa) was a homotetramer with a subunit M(r) of 23 kDa. It contained 1.69 mol of non-heme iron, and had a dark gray color. On anaerobic incubation of it with DHAP, the absorption at around 400 nm increased due to the formation of an enzyme-DHAP complex. Multiple sequence alignment suggested that His77, His79, His115, and Glu96 in the cupin fold were possible metal ligands. The apparent K(m) for DHAP and the apparent V(max) were estimated to be 1.60 μM and 6.28 μmol/min/mg respectively. 2-Hydroxyacetophenone was a poor substrate. CuCl(2) and HgCl(2) strongly inhibited the enzyme, while FeSO(4) weakly activated it.     

Factors contributing to depth perception: behavioral studies on the reverse perspective illusion

In three behavioral experiments using depth-inverted visual stimuli, the factors that contribute to the 'reverse perspective' illusion were measured. The density of linear perspective grid lines was found to induce the illusion most strongly, followed by shading/shadows, and texture/color information. The relative contributions of such pictorial cues to depth perception are similar to those that facilitate the normal perception of 3D space in 2D paintings.     

一个1B/1R小麦-黑麦染色体易位的鉴定

本研究对冬小麦品系73(36)9-1的1B/1R易位染色体进行了遗传分析。发现73(36)9-1有一对随体染色体,它的两个亲本矮丰四号及洛夫林10(Lovrin lo)分别有两对和一对随体染色体。观察用矮丰四号回交的F_1,绝大部分花粉母细胞的中期染色体都能正常配对,而用洛夫林10回交的,除了多数产生两个单价体之外,正常配对的情况也能经常看到。同时还发现,73(36)9-1和“中国春”双端体(CSDT)的1BL能很好地配对并形成一个棒状的异形二价体,而它和CSDT 1BS的染色体则主要产生20″+1′+t′的构型,从而证明易位发生在1B染色体的短臂,并且该易位的片段来自黑麦染色体1RL。本文还讨论了该易位发生的可能途径,推断是由于在F_1花粉母细胞中的两个单价体(一个是小麦染色体1B,一个是黑麦染色体1R)同时进行错分裂之后产生的两种端着丝点染色体(1BL和1RL)重新并合形成的,因而冬小麦73(36)9-1可能是一个自发产生的易位系。     

Isolation and identification of a humanTRPV1 activating compound from soy sauce

Transient receptor potential vanilloid 1 (TRPV1) was identified as a receptor of capsaicin, which is a pungent ingredient in hot red peppers. Due to its relevance for nociception, a physiological and pharmacological study of TRPV1 has also been developed. Therefore, it is important to enrich scientific knowledge regarding the TRPV1 activating or inhibiting compounds. In this study, we fractionated soy sauce based on the human TRPV1 (hTRPV1) activity using column chromatography and purified 5-(9H-pyrido[3,4-b]indol-1-yl)-2-furanmethanol (perlolyrine) as an hTRPV1-activating compound. Additionally, perlolyrine activates the human transient receptor potential ankyrin 1 (hTRPA1). The EC₅₀ of hTRPV1 and hTRPA1 were 2.87 and 1.67 μmol L^?1, respectively. HPLC quantification of soy sauces showed that they contain 2.22–12.13 μmol L^?1 of perlolyrine. The sensory evaluation revealed that perlolyrine has taste modification effect. The results of this study, for the first time, suggest that perlolyrine induces the activation of hTRPV1 and hTRPA1.     

Laminin 511-E8 Fragment Offers Superior Adhesion Properties for Gastric Cancer Cells Compared with Full-Length Laminin 511

Simple SummaryNumerous studies over the past few decades have revealed that the interactions of gastric cancer cells with laminins through integrins play important roles in tumor cell proliferation, infiltration, and metastasis. However, the association between gastric cancer cells and the laminin E8 fragment, which is the smallest integrin-binding component, has not been investigated. In this study, we revealed that the laminin 511-E8 fragment had a greater impact on the adhesion, morphology, and proliferation of gastric cancer cells than full-length laminin 511. Thus, the laminin 511-E8 fragment is considered to be suitable for investigating the interaction between gastric cancer cells and extracellular matrices in tumor invasion and metastasis. Further, the involvement of Cdc42 in the laminin 511-E8 fragment-induced enhanced adhesion of gastric cancer cells was suggested.AbstractBackground: The interaction between cancer cells and laminin (Ln) is a key event in tumor invasion and metastasis. Previously, we determined the effect of full-length Ln511 on gastric cancer cells. However, the interactions between the Ln511-E8 fragment, a truncated protein of Ln511, and gastric cancer cells have not been investigated. Methods: We investigated the adhesion properties of gastric cancer cells to full-length Ln511 and Ln511-E8 fragments. Results: The proliferation of four gastric cancer cell lines (SH-10-TC, MKN74, SC-6-JCK, and MKN45) was highest on the Ln511-E8 fragment. Further, a larger cytoplasm was observed in SH-10-TC and MKN74 cells cultured on full-length Ln511 or Ln511-E8 fragments. The percentage of adhesive cells was highest on the Ln511-E8 fragment in all four cell lines. Moreover, adhesion of the gastric cancer cells to Ln511-E8 fragment-coated plates was reduced by the Cdc42 GTPase inhibitor in a dose-dependent manner, suggesting the involvement of Cdc42 in the Ln511-E8 fragment-induced enhanced adhesion of gastric cancer cells. Conclusions: The Ln511-E8 fragment had a greater impact on the adhesion, morphology, and proliferation of gastric cancer cells than full-length laminin. Thus, the Ln511-E8 fragment is suitable for investigating the interaction between gastric cancer cells and extracellular matrices in tumor invasion and metastasis.     

A 51 kDa Protein Kinase of Potato Activated with Hyphal Wall Components from Phytophthora infestans

In potato tuber tissue, treatment of fungal elicitor, hyphalwall components (HWC) induces various plant defense reactions.As treatment of protein kinase inhibitor prior to HWC treatmentblocks some of defense reactions induced by HWC, involvementof protein kinases in plant defense induction is proposed. Here,we demonstrate HWC-induced activation of a 51-kDa protein kinase(abbreviated p51-PK) using myelin basic protein as a substratein potato tuber discs. The activity of p51-PK was not detectedin the absence of phosphatase inhibitor, NaF, and p51-PK wasimmuroprecipitated with antibody against phosphotyrosine. Pretreatmentof phospholipase C inhibitor, neomycin, and GTP-binding proteinactivator, mastoparan, partially inhibited the HWC-induced activationof p51-PK, suggesting possible involvement of phospholipaseC and GTP-binding protein in the activation of p51-PK. Exogenouslysupplied elicitors, salicylic acid and arachidonic acid, whichare known to induce various defense responses in potato plants,also activated the protein kinase showing the same migrationas p51-PK on SDS-PAGE and different activation patterns. Theseresults implied that p51-PK might be involved in several signaltransduction pathways leading to plant defense responses initiatedby different stimuli. (Received January 11, 1999; Accepted May 25, 1999)     

Requirement of Ala residues at g position in heptad sequence of alpha-helix-forming peptide for formation of fibrous structure

One feature of the alpha3-peptide, which has the amino acid sequence of (Leu-Glu-Thr-Leu-Ala-Lys-Ala)(3), that distinguishes it from many other alpha-helix-forming peptides is its ability to form fibrous assemblies that can be observed by transmission electron microscopy. In this study, the effects of Ala-->Gln substitution at the e (5th) or g (7th) position in the above heptad sequence of the alpha3-peptide on the formation of alpha-helix and fibrous assemblies were investigated by circular dichroism spectral measurement and atomic force microscopy. The 5Qalpha3-peptide obtained by Ala-->Gln substitution at the e position of the alpha3-peptide was found to form very short fibrils with long-elliptical shape, whereas the 7Qalpha3-peptide with Gln residues at the g position lost its ability to form such assemblies, in spite of alpha-helix formation in both peptides; the stabilities of both peptides decreased. These results indicate that Ala residues at the g position in the heptad sequence of the alpha3-peptide are key residues for the formation of fibrous assemblies, which may be due to hydrophobic interactions between alpha-helical bundle surfaces.