New developments for the design,synthesis and biological evaluation of potent SARS-CoV 3CLpro inhibitors

A series of trifluoromethyl, benzothiazolyl or thiazolyl ketone-containing peptidic compounds as SARS-CoV 3CL protease inhibitors were developed and their potency was evaluated by in vitro protease inhibitory assays. Three candidates had encouraging results for the development of new anti-SARS compounds.     

Nucleic Acid Metabolism Involved in Auxin-induced Elongation of Yeast Cells

The following results were obtained using a variant yeast strain, N55, which can respond to the cell-elongating action of auxin. Base analogs of nucleic acids (2-thiouracil, 8-azaguanine, and 5-fluorouracil) inhibited the auxin-induced elongation of yeast cells only when they were added to the preculture prior to auxin treatment. The inhibitory effect of 2-thiouracil and 5-fluorouracil was reversed by uracil and that of 8-azaguanine by guanine. Actino-mycin D inhibited the auxin-induced elongation when given to the culture containing auxin, but not when given to the preculture. The similarity in these respects between yeast and tissues of higher plants is discussed.     

Pulmonary surfactant protein D binds MD-2 through the carbohydrate recognition domain

Pulmonary surfactant protein D (SP-D) is a member of the collectin family and plays crucial roles in the innate immunity of the lung. We have previously shown that surfactant protein A (SP-A), a homologous collectin, interacts with MD-2 and alters lipopolysaccharide signaling. In this study, we examined and characterized the binding of SP-D to MD-2 using a soluble form of recombinant MD-2 (sMD-2). SP-D bound in a concentration- and Ca(2+)-dependent manner to sMD-2 coated onto microtiter wells. Excess mannose abolished the binding of SP-D to sMD-2. In solution, SP-D cosedimented with sMD-2 in the presence of Ca(2+). The direct binding of SP-D to sMD-2 was confirmed by BIAcore analysis. Anti-SP-D monoclonal antibody that recognizes the carbohydrate recognition domain (CRD) of SP-D significantly inhibited the binding of SP-D to sMD-2, indicating the involvement of the CRD for the binding to sMD-2. Ligand blot analysis revealed that SP-D bound to N-glycopeptidase F-treated sMD-2. In addition, the biotinylated SP-D pulled down the mutant sMD-2 with Asn(26) --> Ala and Asn(114) --> Ala substitutions, which lacks the consensus for N-glycosylation. Furthermore, the sMD-2 mutant cosedimented SP-D. These results demonstrate that SP-D directly interacts with MD-2 through the CRD.     

Growth and cell wall changes in azuki bean epicotyls I. Changes in wall polysaccharides during intact growth

Measurement of endogenous growth rates and the mechanical propertyof the cell wall in various regions of light-grown azuki beanepicotyls revealed diat the minimum stress-relaxation time (T_o)was the shortest in the upper region (0–30 mm below theapex) of the epicotyl, where vigorous endogenous growth tookplace, and became longer toward the basal region, which wasmature and not growing. In the upper region of the epicotyl, a lower percentage of a-celluloseand a higher percentage of pectic substances than in the lowerregion were found. The percentage of hemicellulose content wasalmost constant over the whole epicotyl. Major components ofnoncellulosic neutral sugars in the cell wall were galactoseand xylose. The percentage of the galactose content to the noncellulosicpolysaccharide was highest in the upper region and lowest inthe basal region of the epicotyl, and a clearly negative correlationbetween the galactose composition and the T_o value was obtained.On the contrary, the percentage of die xylose content was highestin the basal region and lowest in die upper region, and a clearlypositive correlation between die xylose composition and theT_o value was obtained. During die endogenous growth of die intactepicotyl, all die neutral sugars, particularly galactose, increasedin die upper region, whereas in die middle and basal regions,only xylose increased. Similar changes in sugar compositionswere observed during IAA-induced elongation of die segment excisedfrom various regions of die epicotyl. (Received July 27, 1978; )     

Structural and functional evolution of three cardiac natriuretic peptides

Natriuretic peptides (NPs) are a group of hormones playing important roles in cardiovascular and osmoregulatory systems in vertebrates. Among the NP subtypes, atrial NP (ANP), B-type NP (BNP), and ventricular NP (VNP) are circulating hormones expressed exclusively in the heart (cardiac NPs). The constitution of cardiac NPs is variable among species of vertebrates. In order to understand the evolutionary and functional significance of such variation, we performed a systematic survey of cardiac NP cDNAs in nine taxonomically diverse teleosts inhabiting environments of varying salinity. The discovery of the coexistence of the ANP, BNP, and VNP genes in the eel and rainbow trout suggested that the ancestral teleost had all three cardiac NPs. As the VNP cDNA was undetectable in ayu and six species of Neoteleostei, it is possible that VNP was lost before the divergence of Osmeroidei. The ANP gene was also undetectable in the medaka. Thus, only the BNP gene is universal in species examined in the present study. Synthetic medaka BNP preferentially activated two medaka GC-A-type receptors, suggesting that the three cardiac NPs share the same receptor. However, the regulation of BNP expression may be the most strict because ATTTA repeats in the 3'-untranslated region and the dibasic motif in the ring are conserved among teleosts and tetrapods. Linkage analyses in the rainbow trout located ANP, BNP, and VNP genes on the same chromosome, which suggested the generation of the VNP gene by tandem duplication as observed with ANP and BNP genes. If the duplication occurred before the divergence of tetrapods and teleosts, VNP may exist in the tetrapod lineage.     

Enantioseparation by HPLC using phenylcarbonate, benzoylformate, p-toluenesulfonylcarbamate, and benzoylcarbamates of cellulose and amylose as chiral stationary phases

Phenylcarbonate, benzoylformate, and p-toluenesulfonylcarbamate of cellulose and five new benzoylcarbamate derivatives of both cellulose and amylose were synthesized and their chiral recognition abilities were evaluated as chiral stationary phases (CSPs) for high-performance liquid chromatography (HPLC). Cellulose benzoylcarbamate has a higher chiral recognition ability compared to phenylcarbonate, p-toluenesulfonylcarbamate, and benzoylformate of cellulose. The benzoylcarbamate derivatives exhibited a characteristic chiral recognition for the racemates, which bear a hydrogen atom capable of hydrogen bonding to the carbonyl group of the benzoylcarbamates. The structures of the benzoylcarbamates were investigated by CD spectroscopy.     

Protein kinase C delta regulates Ser46 phosphorylation of p53 tumor suppressor in the apoptotic response to DNA damage

The p53 tumor suppressor is activated in the cellular response to genotoxic stress. Transactivation of p53 target genes dictates cell cycle arrest and DNA repair or induction of apoptosis; however, a molecular mechanism responsible for these distinct functions remains unclear. Recent studies revealed that phosphorylation of p53 on Ser(46) was associated with induction of p53AIP1 expression, resulting in the commitment of the cell fate into apoptotic cell death. Moreover, upon exposure to genotoxic stress, p53DINP1 was expressed and recruited a kinase(s) to p53 that specifically phosphorylated Ser(46). Here, we show that the pro-apoptotic kinase, protein kinase C delta (PKCdelta), is involved in phosphorylation of p53 on Ser(46). PKCdelta-mediated phosphorylation is required for the interaction of PKCdelta with p53. The results also demonstrate that p53DINP1 associates with PKCdelta upon exposure to genotoxic agents. Consistent with these results, PKCdelta potentiates p53-dependent apoptosis by Ser(46) phosphorylation in response to genotoxic stress. These findings indicate that PKCdelta regulates p53 to induce apoptotic cell death in the cellular response to DNA damage.