黑色素是怎样产生的?

答:《生理卫生》教材提到:“生发层中有一些黑色素细胞,能产生黑色素。黑色素含量的多少决定着皮肤颜色的深浅”。在黑色素细胞内苯丙氨酸在苯丙氨酸羟化酶的作用下,首先氧化为酪氨酸、酪氨酸在酪氨酸酶的作用下进一步氧化为二羟基苯丙氨酸(多     

一步法纯化乌骨鸡皮肤酪氨酸酶

酪氨酸酶(EC.1.14.18.1)是黑素细胞内形成黑色素一个关键的酶。它做为一个含铜酶广泛地存在于自然界中。酪氨酸酶既能催化L-酪氨酸的羟化形成L-多巴(羟化酶的活性)又能催化L-多巴的氧化形成多巴醌(氧化酶的活性)。多巴醌进一步自动氧化形成黑色素。     

酪氨酸酶的应用研究进展

酪氨酸酶具有重要的生理生化特性,在医药、环境、食品、精细化工等领域具有广泛的用 途。酪氨酸酶可以氧化L-酪氨酸合成L-多巴和黑色素,L-多巴用于帕金森症的治疗,黑色素能够 杀死HIV病毒。酪氨酸酶可用于环境工程领域处理含苯酚及胺类废水,用于精细化工领域催化 有机合成反应。综述了酪氨酸酶在各个领域的应用概况,阐明了其在工业生产领域的应用前景。     

异色瓢虫酪氨酸羟化酶基因不同发育历期表达及RNAi抑制黑化效果分析

酪氨酸代谢在昆虫黑色素形成中具有重要的调控作用。本研究为探讨酪氨酸代谢调控异色瓢虫黑色素代谢平衡及其发育的影响,先检测异色瓢虫不同发育阶段酪氨酸羟化酶(Tyrosine hydroxylase,TH)基因表达情况,采用RNAi技术对4龄幼虫注射dsTH RNA抑制酪氨酸羟化酶mRNA表达,探索其在蛹期和成虫羽化阶段黑色素形成中的潜在功能。研究结果表明,TH基因在4龄幼虫体内的表达量最高,而在预蛹阶段和羽化后第2天开始表达量显著性下降。其次,dsTH RNA注射后能够显著降低TH本基因的表达,且异色瓢虫4龄幼虫发育缩短,与注射dsGFP RNA组相比较,提前化蛹。最后,dsTH RNA注射后异色瓢虫蛹的重量略有增加,且发育出现异常,即蛹的颜色变化不加深,与对照组相比较,无黑色斑点出现。这些结果表明成虫是在蛹期开始形成黑色素的,及酪氨酸羟化酶能够通过调控酪氨酸代谢途径而影响异色瓢虫化蛹阶段的发育和表皮颜色的变化。相关研究结果有助于丰富酪氨酸代谢调控异色瓢虫表型的内容,为将来保护异色瓢虫,并将其用于害虫生物防治提供理论基础。     

一株高产黑色素细菌的分离及鉴定*

从武汉东湖中分离出一株高产胞外黑色素的细菌(BFHM2002)。BFHM2002具有产黑色素速度快,产量高且不需要酪氨酸诱导等优点;而且经酪氨酸诱导可显提高BF-HM2002胞外黑色素的产量,初步鉴定BFHM2002属坚强芽孢杆菌(Bacillus firmus)。鉴于BFHM2002的产黑色素特性,将成为芽孢杆菌属的一个新的菌种资源。     

内皮素-2促进体外培养的绵羊皮肤黑色素细胞增殖及黑色素生成

内皮素（endothelin,ET）及其受体在黑色素细胞成熟分化时起着有效的促进作用.然而,内皮素-2（ET-2）在黑色素生成的作用方面,还处于争论中或报道不一致.在此,我们研究ET-2对体外培养的绵羊皮肤黑色素细胞增殖和黑色素生成的影响.比较实验组ET-2（1,10,100 nmol/L）与空白对照组,通过MTT法和Ando等的方法检测出黑色素细胞的增殖率和黑色素含量显著增加.荧光定量PCR和Western 印迹分别检测出内皮素受体B(Bdnrb);酪氨酸激酶受体(Kit);酪氨酸酶(Tyr);酪氨酸相关蛋白-1(Tyrp-1)基因的mRNA水平及蛋白水平的相对表达量显著增加.这些数据表明,ET-2可能促进绵羊的皮肤黑色素细胞增殖和黑色素生成.     

酶法合成黑色素的稳定性研究

运用紫外分光光度计对以酪氨酸为底物酶法合成的黑色素进行稳定性分析,结果表明:该黑色素的最大吸收峰为212 nm,不溶于水和酸,易溶于碱,在碱性条件下颜色稳定。热、自然光、紫外光、蔗糖、葡萄糖对色素稳定性无影响;氧化剂H2O2和还原剂Na2SO3对其影响不大;几种金属离子(除Fe2+)对黑色素吸光值及颜色无影响。     

苏云金芽孢杆菌酪氨酸酶基因的克隆表达及应用初探

酪氨酸酶基因编码的酪氨酸酶是生物体合成黑色素的关键酶。采用比较酪氨酸酶的同源保守结构域氨基酸序列的方法设计引物 ,从苏云金芽胞杆菌 (Bacillusthuringiensis) 4D11中通过PCR扩增得到了包含酪氨酸酶基因的DNA片段。将该片段亚克隆到载体pGEM_7zf上并转入大肠杆菌DH5α ,所得到的转化子在添加了L_酪氨酸的LB培养基中能合成可溶性的黑色素。测定该菌株黑色素的产量和在紫外光照射后的菌体活力 ,结果表明该基因产生的黑色素能在一定程度上保护菌体免受紫外辐射     

人工诱导荷包红鲤雌核发育子代与亲本的染色体组型及几种酶的组织化学比较

作者使用元江鲤失活的精子,诱导荷包红鲤卵子进行雌核发育所获得的子代,同亲本作了染色体组型和几种酶的组织化学比较。雌核发育子代的染色体组型与母本荷包红鲤完全相同,与父本元江鲤有不同。元江鲤的体色黑灰色,是由于肌肉中有酪氨酸酶存在,能将酪氨酸氧化成黑色素的缘故;荷包红鲤肌肉中无此酶,不能形成黑色素,所以体色呈桔红色;雌核发育子代肌肉中,也无酪氨酸酶,其体色同母本一样为桔红色;乳酸脱氢酶、非特异性酯酶的分布和含量也都与母本一致,与父本有差异。     

Cortactin tyrosine phosphorylation requires Rac1 activity and association with the cortical actin cytoskeleton

Cortactin is an F-actin binding protein that activates actin-related protein 2/3 complex and is localized within lamellipodia. Cortactin is a substrate for Src and other protein tyrosine kinases involved in cell motility, where its phosphorylation on tyrosines 421, 466, and 482 in the carboxy terminus is required for cell movement and metastasis. In spite of the importance of cortactin tyrosine phosphorylation in cell motility, little is known regarding the structural, spatial, or signaling requirements regulating cortactin tyrosine phosphorylation. Herein, we report that phosphorylation of cortactin tyrosine residues in the carboxy terminus requires the aminoterminal domain and Rac1-mediated localization to the cell periphery. Phosphorylation-specific antibodies directed against tyrosine 421 and 466 were produced to study the regulation and localization of tyrosine phosphorylated cortactin. Phosphorylation of cortactin tyrosine 421 and 466 was elevated in response to Src, epidermal growth factor receptor and Rac1 activation, and tyrosine 421 phosphorylated cortactin localized with F-actin in lamellipodia and podosomes. Cortactin tyrosine phosphorylation is progressive, with tyrosine 421 phosphorylation required for phosphorylation of tyrosine 466. These results indicate that cortactin tyrosine phosphorylation requires Rac1-induced cortactin targeting to cortical actin networks, where it is tyrosine phosphorylated in hierarchical manner that is closely coordinated with its ability to regulate actin dynamics.     

The effect of stress or glucose feeding on hepatic tyrosine aminotransferase activity and liver and plasma tyrosine level of intact and adrenalectomized rats.

The increase of hepatic tyrosine aminotransferase and the fall of plasma tyrosine in rats subjected to immobilization is reconfirmed. Moreover, the same effects three hrs after exposuing the animals to 400 revolutions in Noble-Collip drums are described. However, in bilaterally adrenalectomized rats both hepatic tyrosine aminotransferase and plasma tyrosine remain unchanged after injury and the liver tyrosine level increase. Finally, in animals fed overnight exclusively with 15% glucose solution the well-known decrease of hepatic tyrosine aminotransferase was found paralleled by increased plasma tyrosine levels. A regulatory role of tyrosine aminotransferase in establishing the level of tyrosine in plasma is suggested.     

The contribution of phenylalanine to tyrosine metabolism in vivo. Studies in the post-absorptive and phenylalanine-loaded rat.

1. Rates of appearance and oxidation of plasma L-leucine, L-phenylalanine and L-tyrosine, as well as conversion of plasma phenylalanine into plasma tyrosine, were determined in 90-120 g rats after overnight starvation and while receiving 115-120 mumol of L-phenylalanine/h. 2. In the post-absorptive state, plasma tyrosine and phenylalanine appearances were similar, despite the fact that 22% of plasma tyrosine appearance could be attributed to the hydroxylation of phenylalanine. 3. A constant infusion of 115-120 mumol of L-phenylalanine/h did not significantly alter plasma leucine kinetics, but increased appearance of plasma phenylalanine and tyrosine. The percentage of phenylalanine and tyrosine appearance that was oxidized increased from 12.1% and 24.4% to 37.3% and 48.0% respectively. In phenylalanine-loaded rats, 72% of plasma tyrosine appearance could be attributed to the conversion of phenylalanine. 4. Whole-body tyrosine oxidation measured from a continuous infusion of either L-[14C]tyrosine or L-[14C]phenylalanine differed by 165%. 5. It can be concluded that, in the post-absorptive state, phenylalanine hydroxylation makes a substantial contribution to the plasma appearance of tyrosine and is significantly increased when phenylalanine is administered. The disposal of excess infused phenylalanine is a result of a greater percentage of plasma phenylalanine being converted into tyrosine and a greater proportion of tyrosine being further oxidized. However, apparent tyrosine oxidation rates estimated from plasma tyrosine specific radioactivities and appearance of expired 14CO2 during administration of [14C]tyrosine are underestimates of true rates, in part because tyrosine generated from phenylalanine hydroxylation is catabolized without freely equilibrating with the plasma compartment.     

D‐tyrosine negatively regulates melanin synthesis by competitively inhibiting tyrosinase activity

Although L‐tyrosine is well known for its melanogenic effect, the contribution of D‐tyrosine to melanin synthesis was previously unexplored. Here, we reveal that, unlike L‐tyrosine, D‐tyrosine dose‐dependently reduced the melanin contents of human MNT‐1 melanoma cells and primary human melanocytes. In addition, 500 μM of D‐tyrosine completely inhibited 10 μM L‐tyrosine‐induced melanogenesis, and both in vitro assays and L‐DOPA staining MNT‐1 cells showed that tyrosinase activity is reduced by D‐tyrosine treatment. Thus, D‐tyrosine appears to inhibit L‐tyrosine‐mediated melanogenesis by competitively inhibiting tyrosinase activity. Furthermore, we found that D‐tyrosine inhibited melanogenesis induced by α‐MSH treatment or UV irradiation, which are the most common environmental factors responsible for melanin synthesis. Finally, we confirmed that D‐tyrosine reduced melanin synthesis in the epidermal basal layer of a 3D human skin model. Taken together, these data suggest that D‐tyrosine negatively regulates melanin synthesis by inhibiting tyrosinase activity in melanocyte‐derived cells.     

Tyrosine phosphorylation of the human guanylyl cyclase C receptor

Tyrosine phosphorylation events are key components of several cellular signal transduction pathways. This study describes a novel method for identification of substrates for tyrosine kinases. Co-expression of the tyrosine kinase EphB1 with the intracellular domain of guanylyl cyclase C (GCC) inEscherichia coli cells resulted in tyrosine phosphorylation of GCC, indicating that GCC is a potential substrate for tyrosine kinases. Indeed, GCC expressed in mammalian cells is tyrosine phosphorylated, suggesting that tyrosine phosphorylation may play a role in regulation of GCC signalling. This is the first demonstration of tyrosine phosphorylation of any member of the family of membrane-associated guanylyl cyclases.     

Oncogenic tyrosine kinases target Dok-1 for ubiquitin-mediated proteasomal degradation to promote cell transformation

Cellular transformation induced by oncogenic tyrosine kinases is a multistep process involving activation of growth-promoting signaling pathways and inactivation of suppressor molecules. Dok-1 is an adaptor protein that acts as a negative regulator of tyrosine kinase-initiated signaling and opposes oncogenic tyrosine kinase-mediated cell transformation. Findings that its loss facilitates transformation induced by oncogenic tyrosine kinases suggest that Dok-1 inactivation could constitute an intermediate step in oncogenesis driven by these oncoproteins. However, whether Dok-1 is subject to regulation by oncogenic tyrosine kinases remained unknown. In this study, we show that oncogenic tyrosine kinases, including p210(bcr-abl) and oncogenic forms of Src, downregulate Dok-1 by targeting it for degradation through the ubiquitin-proteasome pathway. This process is dependent on the tyrosine kinase activity of the oncoproteins and is mediated primarily by lysine-dependent polyubiquitination of Dok-1. Importantly, restoration of Dok-1 levels strongly suppresses transformation of cells expressing oncogenic tyrosine kinases, and this suppression is more pronounced in the context of a Dok-1 mutant that is largely refractory to oncogenic tyrosine kinase-induced degradation. Our findings suggest that proteasome-mediated downregulation of Dok-1 is a key mechanism by which oncogenic tyrosine kinases overcome the inhibitory effect of Dok-1 on cellular transformation and tumor progression.     

Formation of tyrosine O-sulfate by mitochondria and chloroplasts of Euglena

Mitochondria that have been purified from cells of light-grown wild-type Euglena gracilis Klebs var. bacillaris Cori or dark-grown mutant W10BSmL and incubated with 35SO4(2-) and ATP accumulate a labeled compound in the surrounding medium. This compound is also labeled when mitochondria are incubated with [14C]tyrosine and nonradioactive sulfate under the same conditions. This compound shows exact coelectrophoresis with synthetic tyrosine O-sulfate at pH 2.0, 5.8, and 8.0, and yields sulfate and tyrosine on acid hydrolysis. Treatment with aryl sulfatase from Aerobacter aerogenes yields sulfate and tyrosine but no tyrosine methyl ester; no hydrolysis of tyrosine methyl ester to tyrosine is observed under identical conditions, ruling out methyl esterase activity in the aryl sulfatase preparation. Thus the compound is identified as tyrosine O-sulfate. No tyrosine O-sulfate is found outside purified developing chloroplasts of Euglena incubated with 35SO4(2-) and ATP, but both chloroplasts and mitochondria accumulate labeled tyrosine-O-sulfate externally when incubated with adenosine 3'-phosphate 5'-phospho[35S]-sulfate (PAP35S). Since tyrosine does not need to be added, it must be provided from endogenous sources. Labeled tyrosine O-sulfate is found in the free pools of light-grown Euglena cells grown on 35SO4(2-) or in dark-grown cells incubated with 35SO4(2-) in light, but none is found in the medium after cell growth. No labeled tyrosine O-sulfate is found in Euglena proteins (including those in extracellular mucus) after growth or incubation of cells with 35SO4(2-) or after incubation of organelles with 35SO4(2-) and ATP or PAP35S, ruling out sulfation of the tyrosine in protein or incorporation of free-pool tyrosine O-sulfate into protein. The system forming tyrosine O-sulfate is membrane-bound and may be involved in transporting tyrosine out of the organelles.     

Inactivation of tyrosine aminotransferase in neutral homogenates and rat liver slices

Inactivation of tyrosine aminotransferase induced in vivo by triamcinolone was studied in a homogenate incubated at neutral pH values. The integrity and the presence of subcellular particles together with a compartment of acidic pH are necessary for inactivation of tyrosine aminotransferase. It is suggested that tyrosine aminotransferase is inactivated inside lysosomes. The system responsible for inactivation of tyrosine aminotransferase was partially purified and identified with lysosomal cathepsins B and B(1). Inactivation of tyrosine aminotransferase in liver slices is controlled by the amino acid concentration and strongly stimulated by cysteine. 3,3',5-Tri-iodo-l-thyronine reversibly and strongly decreases the rate of inactivation of tyrosine aminotransferase. The effect is not due to an increased rate of tyrosine aminotransferase synthesis.     

Identification of the insulin-responsive tyrosine phosphorylation sites on IRSp53

The 53-kDa insulin receptor substrate protein (IRSp53) is part of a regulatory network that organises the actin cytoskeleton in response to stimulation by small GTPases, promoting formation of actin-rich cell protrusions such as filopodia and lamellipodia. It had been established earlier that IRSp53 is tyrosine phosphorylated in response to stimulation of the insulin and insulin-related growth factor receptors, but the consequences of tyrosine phosphorylation for IRSp53 function are unknown. Here, we have used a variety of IRSp53 truncation and point mutants to identify insulin-responsive tyrosine phosphorylation sites on IRSp53. We have found that the C-terminal half of IRSp53 (residues 251-521) undergoes tyrosine phosphorylation in response to insulin stimulation of the insulin beta receptor or epidermal growth factor stimulation via the epidermal growth factor receptor, and that the key residue for insulin receptor-mediated phosphorylation is tyrosine 310, located in a region between the N-terminal IRSp53/MIM homology domain (IMD, residue 1-250) and the central SH3 domain (residues 374-438) that is predicted to be natively unstructured. Mutation of tyrosine 310 to phenylalanine or glutamic acid abrogates the phosphorylation in response to insulin stimulation, but not in response to stimulation of the epidermal growth factor receptor. The N-terminal IMD, which mediates dimerisation of IRSp53, is required for efficient tyrosine phosphorylation downstream of either the insulin or epidermal growth factor receptor stimulation, yet does not appear to include a tyrosine-phosphorylated site itself. Thus, we have identified tyrosine 310 as a primary site of tyrosine phosphorylation in response to insulin signalling and we have shown that although IRSp53 is tyrosine phosphorylated in response to epidermal growth factor receptor signalling, tyrosine 310 is not crucial. Furthermore, the tyrosine phosphorylation status does not appear to affect the cell morphology and production of filopod-like structures upon expression of IRSp53.     

Genome-Wide Analysis and Experimentation of Plant Serine/ Threonine/Tyrosine-Specific Protein Kinases

Protein tyrosine phosphorylation plays an important role in cell growth, development and oncogenesis. No classical protein tyrosine kinase has hitherto been cloned from plants. Does protein tyrosine kinase exist in plants? To address this, we have performed a genomic survey of protein tyrosine kinase motifs in plants using the delineated tyrosine phosphorylation motifs from the animal system. The Arabidopsis thaliana genome encodes 57 different protein kinases that have tyrosine kinase motifs. Animal non-receptor tyrosine kinases, SRC, ABL, LYN, FES, SEK, KIN and RAS have structural relationship with putative plant tyrosine kinases. In an extended analysis, animal receptor and non-receptor kinases, Raf and Ras kinases, mixed lineage kinases and plant serine/threonine/tyrosine (STY) protein kinases, form a well-supported group sharing a common origin within the superfamily of STY kinases. We report that plants lack bona fide tyrosine kinases, which raise an intriguing possibility that tyrosine phosphorylation is carried out by dual-specificity STY protein kinases in plants. The distribution pattern of STY protein kinase families on Arabidopsis chromosomes indicates that this gene family is partly a consequence of duplication and reshuffling of the Arabidopsis genome and of the generation of tandem repeats. Genome-wide analysis is supported by the functional expression and characterization of At2g24360 and phosphoproteomics of Arabidopsis. Evidence for tyrosine phosphorylated proteins is provided by alkaline hydrolysis, anti-phosphotyrosine immunoblotting, phosphoamino acid analysis and peptide mass fingerprinting. These results report the first comprehensive survey of genome-wide and tyrosine phosphoproteome analysis of plant STY protein kinases.