细菌蛋白质磷酸化修饰研究进展

细菌蛋白质磷酸化修饰是调控细菌基因表达的一种重要方式,在细菌诸多生命活动中发挥非常关键的作用。本文系统概括了近年来细菌蛋白质磷酸化修饰的种类、双组分调控系统中磷酸化修饰调控信号传导、酪氨酸残基磷酸化修饰以及丝/苏氨酸残基磷酸化修饰等,同时对不同种类细菌蛋白质磷酸化修饰的功能进行综述,这些研究将对人类了解细菌蛋白质翻译后修饰的磷酸化调控及其与控制细菌感染的关系提供参考价值。     

突触可塑性调节过程中蛋白质磷酸化修饰的研究进展

突触可塑性可以导致神经元传递效率的改变,是神经系统发育、学习记忆等脑的高级功能活动中细胞功能的重要基础。蛋白质磷酸化修饰通过蛋白激酶和蛋白磷酸酶之间的动态平衡对突触可塑性和突触传递的长期调节,参与各种脑疾病(包括精神疾病和神经退行性疾病)的发生发展。本文综述了磷酸化修饰和突触可塑性的关系,重点介绍了长时程增强和长时程抑制相关的离子型谷氨酸受体磷酸化修饰研究进展,以期为神经元突触可塑性改变相关的脑疾病研究提供新的思路。     

杨树蛋白质磷酸化位点预测

以小黑杨磷酸化蛋白质组为研究对象,用人工神经网络表达丝氨酸、苏氨酸等残基位点的磷酸化与氨基酸序列的结构特征之间的非线性关系,建立了BP人工神经网络模型,并用磷酸化数据对所建模型进行训练和分析,得适宜的结构为21×16∶8∶4,拟合准确度为90%,Acc、Sn、Sp、MCC分别为78%、89%、67%、0.57,对比分析结果表明,所建模型具有较强的预测能力。     

光色素信号通路中磷酸化修饰研究进展

光是植物的唯一能量来源, 植物在进化过程中产生不同的光敏色素来感知光信号。光信号通路中元件通常被特异翻译后修饰调节。光敏色素是一种自磷酸化的丝氨酸/苏氨酸蛋白激酶, 可以被一些蛋白磷酸酶去磷酸化。通过对光敏色素A (phyA)和光敏色素B (phyB)的自磷酸化位点研究, 发现自磷酸化对光敏色素的功能及其介导的信号通路起着非常重要的作用。光激活的光敏色素诱导光敏色素作用因子(PIF)磷酸化, 这对于PIF的正常降解及光形态建成的起始是必需的。该文主要介绍了光敏色素信号通路磷酸化修饰的最新进展, 以期为深入研究光敏色素信号转导机制提供参考。     

蛋白质O-GlcNAc糖基化修饰对tau蛋白磷酸化修饰的影响

蛋白质的O位N-乙酰葡萄糖胺(O-GlcNAc)糖基化修饰是一种新近发现的广泛存在于细胞核蛋白与细胞浆蛋白的蛋白质翻译后修饰.其性质与经典的膜蛋白和分泌蛋白的糖基化修饰不同,而与蛋白质磷酸化修饰更相似.O-GlcNAc糖基化和磷酸化均修饰tau蛋白的丝氨酸和苏氨酸残基,通过改变O-GlcNAc糖基化供体底物浓度以及其关键酶活性等方法,改变分化后成神经细胞样的PC12细胞中的蛋白质O-GlcNAc糖基化修饰水平,然后用特异性识别不同位点磷酸化的tau蛋白抗体,进行蛋白质印迹分析来检测tau蛋白磷酸化水平的变化.结果发现细胞内蛋白质O-GlcNAc糖基化对tau蛋白磷酸化的影响,在不同的磷酸化位点其影响不同.增加蛋白质O-GlcNAc糖基化修饰导致tau蛋白大多数磷酸位点的磷酸化水平降低,反之亦然.这些结果说明,tau磷酸化在大多数位点受到O-GlcNAc糖基化修饰的负性调节.这一研究为阐明调节tau蛋白磷酸化水平的机理和阿尔茨海默病脑中tau异常过度磷酸化的分子机制提供了新的线索.     

植物蛋白质磷酸化的研究技术

本文介绍植物蛋白质磷酸化研究的技术及其应用情况,并对这些技术的应用前景作了展望。     

抗病毒核苷酸类似物磷酸化修饰研究进展

核苷(酸)类似物是一类抗病毒前药,其进入人体细胞后经过逐步磷酸化生成核苷三磷酸类似物发挥抗代谢药作用,主要通过抑制病毒复制和促进侵染细胞凋亡,达到疾病治疗效果.其中,核苷类似物在细胞内经激酶活化的代谢转化过程通常是不充分的,导致最后生成的核苷三磷酸类似物浓度较低,降低了作用效果.因此,通过直接制备核苷酸类似物作为抗病毒...     

蛋白磷酸化修饰在植物-病原微生物互作中的作用研究进展

蛋白磷酸化修饰是植物细胞信号调控的普遍机制。植物-病原微生物互作过程中, 关键调控蛋白的磷酸化状态影响免疫信号的激活。多种病原微生物通过干扰宿主蛋白的磷酸化状态攻击免疫系统, 以提高致病性。该文对植物免疫调控过程中关键元件的磷酸化修饰及其在免疫信号中的调控作用进行了综述。研究植物-病原菌互作过程中关键蛋白的磷酸化修饰, 有助于深入探讨植物-病原微生物互作的分子机理。该文将为寻找广谱抗病的新途径提供理论依据。     

蛋白质翻译后修饰研究进展

翻译后修饰在蛋白质加工、成熟的过程中发挥着重要的作用,它可以改变蛋白质的物理、化学性质,影响蛋白质的空间构象、立体位阻及其稳定性,进而对蛋白质的生物学活性产生作用,引起蛋白质的功能改变。修饰基团自身的结构特性对蛋白质的性质、功能也会产生深远的影响。在已有的研究基础上,综述蛋白质翻译后修饰的主要类型以及各修饰作用潜在的生物学功能。     

HIV-1 TAT蛋白转导肽的研究进展

TAT蛋白转导肽是人类免疫缺陷病毒1型(human immunodeficiency virus type 1, HIV-1)编码的一段富含碱性氨基酸、带正电荷的多肽,属于蛋白转导域家族的一员。长期研究发现其全长及11个碱性氨基酸富集区的核心肽段(YGRKKRRQRRR)不仅能够在包括蛋白质、多肽及核酸等多种外源生物大分子的跨膜转导过程中具有重要作用,而且能够携带这些外源生物大分子通过活体细胞的各种生物膜性结构(如细胞膜和血脑屏障等)并发挥生理功能,但其跨膜转导机制仍不明确。新近研究还发现TAT核心肽段在促进外源蛋白高效表达过程中也具有重要作用,能够显著增加外源蛋白高效、可溶性表达的水平,显示了TAT蛋白转导肽的新功能。以TAT蛋白转导肽跨膜转导作用的长期研究背景为基础,分别从TAT蛋白转导肽的结构特点、其跨膜转导作用的影响因素及其作用机制等方面进行了系统综述,进一步结合TAT蛋白转导肽的最新研究进展分别从药物研发、机制探索及新功能的开发等方面展望了后续研究方向与应用价值,不仅为深入阐述TAT蛋白转导肽的跨膜转导作用的功能意义提供了参考依据,而且为TAT蛋白转导肽在微生物工程及蛋白质工程等领域的潜在应用价值提供了重要参考信息。     

An Exploration of the Effects of Constraints on the Phosphorylation of Synthetic Protein Tyrosine Kinase Peptide Substrates

We synthesized by classical solution methods three conformational constrained analogues of EDNEYTA, a heptapeptide sequence that represents the common major autophosphorylation site of the protein tyrosine kinases (PTKs) of the Src family. The correlation between the different structural properties induced by the modifications of the native sequence and the propensity of the peptides to act as PTK substrates was examined. The kinetic data obtained indicate that the introduction of the tyrosine-analogue constraints Tic(OH) and MeTyr, which block the ring flexibility, completely prevents the phosphorylation catalysed by the kinases Lyn and Fgr. On the other hand PTKIIB/p38^syk can phosphorylate the two derivatives albeit with an efficiency lower than that found with the native sequence. A third derivative contained side chain to side chain cyclization. This analogue, in which the freedom of the phenolic moiety is not altered, can be phosphorylated by all the PTKs tested with kinetic constants comparable to the parent peptide.     

Protein Ser/Thr/Tyr Phosphorylation in the Archaea

The third domain of life, the Archaea (formerly Archaebacteria), is populated by a physiologically diverse set of microorganisms, many of which reside at the ecological extremes of our global environment. Although ostensibly prokaryotic in morphology, the Archaea share much closer evolutionary ties with the Eukarya than with the superficially more similar Bacteria. Initial genomic, proteomic, and biochemical analyses have revealed the presence of “eukaryotic” protein kinases and phosphatases and an intriguing set of serine-, threonine-, and tyrosine-phosphorylated proteins in the Archaea that may offer new insights into this important regulatory mechanism.     

毛细管区带电泳／串联质谱联用法鉴定多肽和蛋白质

建立了毛细管区带电泳－串联质谱联用（ＣＺＥ／ＭＳ／ＭＳ）对多肽和蛋白质高灵敏度鉴定方法,对Ｍｅｔ－脑啡肽和Ｌｅｕ－脑啡肽的混合物进行了分析,用ＣＺＥ／ＭＳ／ＭＳ方法验证了各自的序列,同样对细胞色素ｃ的胰蛋白酶酶解产物用ＣＺＥ／ＭＳ／ＭＳ方法进行了肽质谱分析,几科所有肽段的序列及其与在分子中的位置都得到了确定,通过ＳＥＱＵＥＳＴ软件进行蛋白质序列数据库搜索得到准确的鉴定结果,所消耗的样品量均在低皮可     

Protein Phosphorylation and APP Metabolism

Numerous lines of evidence place signal transduction cascades at the core of many processes having a direct role in neurodegeneration and associated disorders. Key players include neurotransmitters, growth factors, cytokines, hormones, and even binding and targeting proteins. Indeed, abnormal phosphorylation of key control proteins has been detected in many cases and is thought to underlie the associated cellular dysfunctions. Several signaling cascades have been implicated, affecting processes as varied as protein processing, protein expression, and subcellular protein localization, among others. The Alzheimer's amyloid precursor protein (APP) is a phosphoprotein, with well-defined phosphorylation sites but whose function is not clearly understood. The factors and pathways regulating the processing of APP have been particularly elusive, both in normal ageing and the Alzheimer's disease (AD) condition. Not surprisingly, the physiological function(s) of the protein remain(s) to be elucidated, although many hypotheses have been advanced. Nonetheless, considerable data has accumulated over the last decade, placing APP in key positions to be modulated both directly and indirectly by phosphorylation and phosphorylation-dependent events. The pathological end product of APP processing is the main proteinaceous component of the hallmark senile plaques found in the brains of AD patients, that is, a toxic peptide termed A. In this minireview we address the importance of phosphorylation and signal transduction cascades in relation to APP processing and A production. The possible use of the identified molecular alterations as therapeutic targets is also addressed.     

Protein Phosphorylation in Astrocytes Mediated by Protein Kinase C: Comparison with Phosphorylation by Cyclic AMP-Dependent Protein Kinase

The protein kinase C activator, phorbol 12-myristate 13-acetate (PMA), has been found recently to transform cultured astrocytes from flat, polygonal cells into stellate-shaped, process-bearing cells. Studies were conducted to determine the effect of PMA on protein phosphorylation in astrocytes and to compare this pattern of phosphorylation with that elicited by dibutyryl cyclic AMP (dbcAMP), an activator of the cyclic AMP-dependent protein kinase which also affects astrocyte morphology. Exposure to PMA increased the amount of 32P incorporation into several phosphoproteins, including two cytosolic proteins with molecular weights of 30,000 (pI 5.5 and 5.7), an acidic 80,000 molecular weight protein (pI 4.5) present in both the cytosolic and membrane fractions, and two cytoskeletal proteins with molecular weights of 60,000 (pI 5.3) and 55,000 (pI 5.6), identified as vimentin and glial fibrillary acidic protein, respectively. Effects of PMA on protein phosphorylation were not observed in cells depleted of protein kinase C. In contrast to the effect observed with PMA, treatment with dbcAMP decreased the amount of 32P incorporation into the 80,000 protein. Like PMA, treatment with dbcAMP increased the 32P incorporation into the proteins with molecular weights of 60,000, 55,000 and 30,000, although the magnitude of this effect was different. The effect of dbcAMP on protein phosphorylation was still observed in cells depleted of protein kinase C. The results suggest that PMA, via the activation of protein kinase C, can alter the phosphorylation of a number of proteins in astrocytes, and some of these same phosphoproteins are also phosphorylated by the cyclic AMP-dependent mechanisms.     

Regiospecific Phosphorylation Control of the SR Protein ASF/SF2 by SRPK1

SR proteins (splicing factors containing arginine-serine repeats) are essential factors that control the splicing of precursor mRNA by regulating multiple steps in spliceosome development. The prototypical SR protein ASF/SF2 (human alternative splicing factor) contains two N-terminal RNA recognition motifs (RRMs) (RRM1 and RRM2) and a 50-residue C-terminal RS (arginine-serine-rich) domain that can be phosphorylated at numerous serines by the protein kinase SR-specific protein kinase (SRPK) 1. The RS domain [C-terminal domain that is rich in arginine-serine repeats (residues 198-248)] is further divided into N-terminal [RS1: N-terminal portion of the RS domain (residues 198-227)] and C-terminal [RS2: C-terminal portion of the RS domain (residues 228-248)] segments whose modification guides the nuclear localization of ASF/SF2. While previous studies revealed that SRPK1 phosphorylates RS1, regiospecific and temporal-specific control within the largely redundant RS domain is not well understood. To address this issue, we performed engineered footprinting and single-turnover experiments to determine where and how SRPK1 initiates phosphorylation within the RS domain. The data show that local sequence elements in the RS domain control the strong kinetic preference for RS1 phosphorylation. SRPK1 initiates phosphorylation in a small region of serines (initiation box) in the middle of the RS domain at the C-terminal end of RS1 and then proceeds in an N-terminal direction. This initiation process requires both a viable docking groove in the large lobe of SRPK1 and one RRM (RRM2) on the N-terminal flank of the RS domain. Thus, while local RS/SR content steers regional preferences in the RS domain, distal contacts with SRPK1 guide initiation and directional phosphorylation within these regions.