Induction of chalcone synthase in cell suspension cultures of carrot (Daucus carota L. spp. sativus) by ultraviolet light: evidence for two different forms of chalcone synthase

Two cell lines of carrot (Daucus carota L. spp. sativus), grown as cell-suspension cultures in the dark, were irradiated with ultraviolet light (315–420 nm) 10 d after the onset of cultivation. Chalcone synthase (CHS) enzyme activity was induced in both cell lines. Anthocyanin synthesis was only stimulated in the anthocyanin-containing cell line DCb. Parallel to the increase in CHS activity there was an increase with time in the amount of one CHS form with an isoelectric point of 6.5 and a molecular weight of 40 kilodaltons (kDa) per subunit. Whereas the anthocyanin-free cell line DCs failed to accumulate anthocyanin, it did stimulate another CHS form with an isoelectric point at pH 5.5 and a molecular weight of 43 kDa per subunit. Both enzyme activities could be separated by isoelectric focusing and stabilized using sodium hydrosulfite as an oxidation protectant. In carrot plants, CHS was restricted to the dark purple petals of the inflorescence (40 kDa) and to the leaves (43 kDa).Abbreviations BSA bovine serum albumin - CHS chalcone synthase - IEF isoelectric focusing - kDa kilodaltons - KP_i potassium phosphate buffer - PAL phenylalanine ammonialyase - pI isoelectric point - UV ultraviolet     

一种作用于花青素和甜菊醇的甜菊糖基转移酶的基因克隆和功能分析

本文对一种新的甜菊糖基转移酶进行了基因克隆和功能分析。获得的基因cDNA全长1419bp,编码473个氨基酸,蛋白质分子量约53.2K Da。与常见的糖基转移酶基因比较,相似性达44%以上,且具有糖基转移酶的保守序列。体外异源表达获得的融合蛋白,具有在花青素类和甜菊醇等糖基受体上转糖基的酶活性。在对一系列不同底物的酶活性进行比较后,推测这种糖基转移酶在体内参与了甜菊糖苷的合成。结果表明,具有广泛的底物活性的类黄酮类糖基转移酶,在甜菊体内不仅对类黄酮转糖基,而且在生成水溶性甜菊糖苷的过程中也扮演重要的角色。     

5种野牡丹属植物花色素成分及影响花色呈色因子分析

为明确野牡丹属(Melastoma L.)植物花瓣的色素成分和呈色机理,为花色育种提供参考。以野牡丹(M.candidum)、白花野牡丹(M.candidum f.albiflorum)、印度野牡丹(M.malabathiricum)、白花印度野牡丹(M. malabathricumvar.alba)、毛稔(M.sanguinrum)5种野牡丹属植物材料,采用目测法、RHSCC比色法和色差仪测定花瓣表型,应用化学显色法、紫外分光光度法对花色素成分及含量进行初步分析与测定,通过徒手切片组织切片法观察花瓣表皮细胞的显微结构和分布特点,测定花瓣pH值、可溶性糖及可溶性蛋白含量等生理指标分析对花色的影响。结果显示,野牡丹属植物花瓣不含叶绿素和类胡萝卜素,紫罗兰色系主要含花青素苷和黄酮类化合物,白色系主要含黄酮类化合物。野牡丹和毛稔花色素分布于上、下表皮,印度野牡丹花色素分布于上、下表皮和栅栏组织,白花野牡丹和白花印度野牡丹花瓣没有发现色素积累;紫罗兰色系野牡丹上表皮细胞呈圆锥形突起,白色系野牡丹上表皮细胞呈不规则的扁平状,它们下表皮细胞全呈不规则的扁平状。野牡丹属植物花色明度L*随花瓣颜色变深而降低,明度L*与红度a*呈极显著负相关、与蓝度b*呈极显著的正相关。花瓣中花青素苷含量与其明度L*和蓝度b*呈显著负相关,pH值与花瓣红度a*呈现显著的负相关。研究表明,野牡丹属植物花色主要由花青素苷决定,花青素苷含量、色素分布、上表皮细胞形状等是引起花色呈现多样的主要因子。     

Uridine 5′-diphosphate-xylose: anthocyanidin 3-O-glucose-xylosyltransferase from petals of Matthiola incana R.Br.

Petals of genetically defined lines of Matthiola incana R.Br. contain a glycosyltransferase which catalyzes the transfer of the xylosyl moiety of uridine 5-diphosphate-xylose to the glucose of cyanidin 3-glucoside. The enzyme also uses 3-glucosides of pelargonidin and delphinidin, cyanidin 3-(p-coumaroyl)-glucoside and 3-(caffeoyl)-glucoside as substrates. The xylosyltransferase exhibits a pH optimum of 6.5. The enzyme activity depends on the stage of bud and flower development. Accumulation of cyanidin 3-glucoside during flower development is correlated with xylosyltransferase activity.Abbreviations HPLC high-performance liquid chromatography - UDP uridine 5-diphosphate     

Flavonoid methylation: a novel 4'-O-methyltransferase from Catharanthus roseus, and evidence that partially methylated flavanones are substrates of four different flavonoid dioxygenases

Catharanthus roseus (Madagascar periwinkle) flavonoids have a simple methylation pattern. Characteristic are B-ring 5' and 3' methylations and a methylation in the position 7 of the A-ring. The first two can be explained by a previously identified unusual O-methyltransferase (CrOMT2) that performs two sequential methylations. We used a homology based RT-PCR strategy to search for cDNAs encoding the enzyme for the A-ring 7 position. Full-length cDNAs for three proteins were characterized (CrOMT5, CrOMT6, CrOMT7). The deduced polypeptides shared 59-66% identity among each other, with CrOMT2, and with CrOMT4 (a previously characterized protein of unknown function). The five proteins formed a cluster separate from all other OMTs in a relationship tree. Analysis of the genes showed that all C. roseus OMTs had a single intron in a conserved position, and a survey of OMT genes in other plants revealed that this intron was highly conserved in evolution. The three cDNAs were cloned for expression of His-tagged recombinant proteins. CrOMT5 was insoluble, but CrOMT6 and CrOMT7 could be purified by affinity chromatography. CrOMT7 was inactive with all compounds tested. The only substrates found for CrOMT6 were 3'-O-methyl-eriodictyol (homoeriodictyol) and the corresponding flavones and flavonols. The mass spectrometric analysis showed that the enzyme was not the expected 7OMT, but a B-ring 4'OMT. OMTs with this specificity had not been described before, and 3',4'-dimethylated flavonoids had not been found so far in C. roseus, but they are well-known from other plants. The identification of this enzyme activity raised the question whether methylation could be a part of the mechanisms channeling flavonoid biosynthesis. We investigated four purified recombinant 2-oxoglutarate-dependent flavonoid dioxygenases: flavanone 3beta-hydroxylase, flavone synthase, flavonol synthase, and anthocyanidin synthase. 3'-O-Methyl-eriodictyol was a substrate for all four enzymes. The activities were only slightly lower than with the standard substrate naringenin, and in some cases much higher than with eriodictyol. Methylation in the A-ring, however, strongly reduced or abolished the activities with all four enzymes. The results suggested that B-ring 3' methylation is no hindrance for flavonoid dioxygenases. These results characterized a new type of flavonoid O-methyltransferase, and also provided new insights into the catalytic capacities of key dioxygenases in flavonoid biosynthesis.     

11个红紫芽茶树新品系的芽叶特性和生化成分研究

对全年芽叶都呈红紫色的11个新选育的高花青素茶树品系新梢芽叶进行了百芽重和色泽调查,并对其生化组分进行了比较分析,旨在为今后筛选出高产优质的高花青素茶树种质资源、开发高花青素茶叶加工和深加工保健产品提供物质和理论基础。结果表明:11个新品系分枝能力较强,百芽重适中,具有较高的产量潜力;芽叶红紫色的深浅与花青素含量呈正相关,而与茶多酚、儿茶素、咖啡碱和游离氨基酸无关;除HY-5和HY-8芽叶色泽在秋季最深外,其他红紫芽品系芽叶的红紫色都在夏季最深,说明红紫芽茶树芽叶的色泽及其花青素含量同时受外界环境和遗传因子的控制。