Characterization of the side-chain hydroxyl moieties of residues Y56, Y111, Y238, Y338, and S339 as determinants of specificity in E. coli cystathionine β-lyase

Cystathionine β-lyase (CBL) catalyzes the hydrolysis of L-cystathionine (L-Cth) to produce L-homocysteine, pyruvate, and ammonia. A series of site-directed variants of Escherichia coli CBL (eCBL) was constructed to investigate the roles of the hydroxyl moieties of active-site residues Y56, Y111, Y238, Y338, and S339 as determinants of specificity. The effect of these conservative substitutions on the k(cat)/K(m)(L-Cth) for the α,β-elimination of L-Cth ranges from a change of only 1.1-fold for Y338F to a reduction of 3 orders of magnitude for the alanine replacement variant of S339. A novel role for residue S339 as a determinant of reaction specificity, via tethering of the catalytic base, K210, is demonstrated. Comparison of the kinetic parameters for L-Cth hydrolysis with those for the inhibition of eCBL by aminoethoxyvinylglycine (AVG) indicates that Y238 interacts with the distal carboxylate group of the substrate. The 22 and 50-fold increases in the K(m)(L-Cth) and K(i)(AVG) resulting from replacement of Y56 with phenylalanine suggest that this residue may interact with the distal amino group of these compounds, although an indirect role in binding is more likely. The near-native k(cat)/K(m)(L-Cth) and pH profile of the eCBL-Y111F variant demonstrate that residue Y111 does not play a role in proton transfer. The understanding of the eCBL active site and of the determinants of substrate and reaction specificity resulting from this work will facilitate the design of inhibitors, as antibacterial therapeutics, and the engineering of enzymes dependent on the catalytically versatile pyridoxal 5'-phosphate cofactor to modify reaction specificity.     

A Neglected Character of Cornus L. s. 1. with Special Reference to a New Subgenus—Sinocornus Q. Y. Xiang

Cornus L. s. 1. is a large polytypic genus. The classificantion of which has been so different that some authors recognize several separate genera, while others treat them as either subgenera or sections. New evidence from many disciplines such as palynology, cytology, wood anatomy and embryology supports the view that the genus should remain in the broad sense. I basically agree with the treatment by Ferguson (1966_b) before my finishing a comprehensive study on Cornus L. s. 1. except for supporting the separation of subgenus Afrocrania as an independent genus. A new subgenus Sinocornus Q. Y. Xiang, containing one species, C. chinensis Wangerin, is established here on account of its inflorescence axillary and different from the other members of the genus which all have terminal inflorescences only. The inflorescence developed from a terminal bud implicates the sympodial nature of axis of stem and the one from an axillary bud the monopodial one. It is reasonable to regard the character as of subgeneric value. Also a new combination, Cornus subgen. Syncarpea (Nakai) Q. Y. Xiang, is proposed here as a substitute for Cornus subgen. Benthamia (C. B. Clarke) Schneider used for a long time. Because the valid publication of the former name subgen. Syncarpea Nakai is earlier than the latter one. The types of inflorescences of 8 subgenera represented by type or selected species are illustrated.     

Virulence as a consequence of genome instability of a novel temperate bacteriophage, ΦRsG1, of Rhodobacter sphaeroides Y

A novel temperate bacteriophage, designated RsG1, was isolated from Rhodobacter sphaeroides Y (previously designated Rhodopseudomonas sphaeroides) following exposure to mitomycin C. The phage morphology, as revealed from electron microscopy, showed a hexagonal head (90 by 46.5 nm) connected with a tail (116 by 9.4 nm), to which a collar was proximally attached. A morphologically similar phage was also produced by spontaneous lysis of the cells. While RsG1 did not grow on any other bacterial strain tested, spontaneously produced phage particles propagated (and formed plaques) on R. sphaeroides Y still carrying RsG1 in the prophage state. The genome of RsG1 consisted of double stranded linear DNA with cohesive ends and a GC-content of 71.8 mol%. The DNA molecules formed circles in vitro with a mean contour length of 17.18±0.4 m, which corresponds to a size of 49 kbase pairs (kb). On the other hand, DNA extracted from the virulent phage particles was heterogeneous and consisted of two DNA species of different size, occurring in a ratio of about 1:1. These molecules also circularized having contour lengths of 17.18±0.4 m and 14.02±0.41 m corresponding to 49 and 40 kb, respectively. Restriction digest analysis of the two DNA species and DNA from RsG1 indicated that they are similar, and allowed the indentification of an 11.5 kb EcoRI fragment that carries the cohesive ends. Because DNA from RsG1 and the 49 kb DNA of the virulent phage particles were indistinguishable with the criteria applied, it is suggested that phage particles containing the 40 kb DNA represent the virulent type of phage, termed RsG1.1.     

Reappraisal of Microtropis macrocarpa C.Y. Cheng ＆ T.C. Kao （Celastraceae）, a critically endangered species endemic to Yunnan,China

Microtropis macrocarpa C.Y. Cheng ＆ T.C. Kao has been treated as a synonym of Microtropis macrophylla Merr. ＆ Freeman in most taxonomic reports in China. According to our study, M. macrocarpa is an independent species endemic to Yunnan, China. Microtropis macrophylla and Microtropis pachyphylla Merr. ＆ Freeman should be treated as synonyms of Microtropis longifolia Wall. ex Kurz. M. macrocarpa is also identified as a critically endangered species, CR B lab （i,ii,iii,v）, based on the IUCN Red List Categories and Criteria. The foliar surface, anther and pollen structures were also observed with a scanning electron microscope.     

β-Carboline alkaloids from the leaves of Trigonostemon lii Y.T. Chang

A phytochemical work on the alkaloid constituents from Trigonostemon lii Y.T. Chang was conducted to give six new β-carboline alkaloids, trigonostemines A-F (1-6) and eight known β-carboline alkaloids (7-14). Their structures were elucidated by extensive spectroscopic techniques including 2D NMR experiments and mass spectrometry. All of the compounds were evaluated for their cytotoxic activities against the HL-60, SMMC-7721, A-549, MCF-7, and SW480 human cancer cell lines. Trigonostemines A and B (1 and 2) exhibited stronger inhibitory activities than the positive control (cisplatin) in some cell lines.