Crystal structure of a protein phosphatase 2A heterotrimeric holoenzyme

Protein phosphatase 2A (PP2A) is a principal Ser/Thr phosphatase, the deregulation of which is associated with multiple human cancers, Alzheimer's disease and increased susceptibility to pathogen infections. How PP2A is structurally organized and functionally regulated remains unclear. Here we report the crystal structure of an AB'C heterotrimeric PP2A holoenzyme. The structure reveals that the HEAT repeats of the scaffold A subunit form a horseshoe-shaped fold, holding the catalytic C and regulatory B' subunits together on the same side. The regulatory B' subunit forms pseudo-HEAT repeats and interacts with the C subunit near the active site, thereby defining substrate specificity. The methylated carboxy-terminal tail of the C subunit interacts with a highly negatively charged region at the interface between A and B' subunits, suggesting that the C-terminal carboxyl methylation of the C subunit promotes B' subunit recruitment by neutralizing charge repulsion. Together, our structural results establish a crucial foundation for understanding PP2A assembly, substrate recruitment and regulation.     

A protein-tyrosine phosphatase with sequence similarity to the SH2 domain of the protein-tyrosine kinases.

The phosphorylation of proteins at tyrosine residues is critical in cellular signal transduction, neoplastic transformation and control of the mitotic cycle. These mechanisms are regulated by the activities of both protein-tyrosine kinases (PTKs) and protein-tyrosine phosphatases (PTPases). As in the PTKs, there are two classes of PTPases: membrane associated, receptor-like enzymes and soluble proteins. Here we report the isolation of a complementary DNA clone encoding a new form of soluble PTPase, PTP1C. The enzyme possesses a large noncatalytic region at the N terminus which unexpectedly contains two adjacent copies of the Src homology region 2 (the SH2 domain) found in various nonreceptor PTKs and other cytoplasmic signalling proteins. As with other SH2 sequences, the SH2 domains of PTP1C formed high-affinity complexes with the activated epidermal growth factor receptor and other phosphotyrosine-containing proteins. These results suggest that the SH2 regions in PTP1C may interact with other cellular components to modulate its own phosphatase activity against interacting substrates. PTPase activity may thus directly link growth factor receptors and other signalling proteins through protein-tyrosine phosphorylation.     

A Tyr/Ser protein phosphatase encoded by vaccinia virus.

Protein tyrosine phosphorylation is associated with alterations in receptor activity, cellular proliferation and modulation of the cell cycle. Inappropriate tyrosine phosphorylation can lead to unrestrained cell growth and oncogenesis. Enzymes important in tyrosine dephosphorylation have also been described. Protein tyrosine phosphatases (PTPases) consist of two families. There is a receptor-like family of PTPases with an extracellular domain, transmembrane-spanning region and typically two repeated phosphatase domains. Proteins of the non-receptor-like family have a single catalytic phosphatase domain, show a substrate specificity for Tyr phosphate and will not hydrolyse Ser or Thr phosphate. Here we report that the vaccinia virus genome contains an open reading frame which shares amino-acid sequence identity with the PTPases. The purified protein encoded by the vaccinia virus H1 open reading frame expressed in bacteria hydrolyses substrates containing phosphotyrosine and phosphoserine. Mutagenesis of an essential Cys in the vaccinia phosphatase abolishes catalytic activity directed towards both substrates, suggesting that hydrolysis proceeds by a common mechanism. Understanding the function of the H1-encoded protein will help to define the role of the phosphatase in viral replication and pathogenesis.     

苦荞蛋白磷酸酶2C家族的鉴定及表达分析

为了研究苦荞蛋白磷酸酶2C(PP2C)家族的成员和分类,及后续探究其在苦荞生长发育中的功能,本文利用生物信息学方法对苦荞PP2C家族进行鉴定、分类,并对其基因结构、保守基序、分子进化等进行分析.结果表明,苦荞PP2C家族有81个成员,划分为A-K的11个亚族,并且在同亚族中序列特征相似,而不同亚族间序列特征有一定差异;苦荞PP2C家族有14次基因重复事件.此外,qRT-PCR分析结果表明其A亚族基因在苦荞根、茎、叶、花、果中均有表达,除FtPP2C44外的8个基因在苦荞花、果中表达量较高;在苦荞幼苗中,除FtPP2C08外的8个基因均受ABA诱导表达量上调.上述结果揭示了苦荞PP2C家族的成员组成、序列特征、扩增和其A亚族基因的组织表达模式及受ABA诱导表达情况.     

A Fingerprinting Method Based on Module Extraction Used to Protect Intellectual Property

Intellectual Property （IP） reuse methodology has been widely used in Integrate Circuit （IC） design. Meanwhile, the corresponding security problems caused by illegal IP distribution have aroused lots of attentions. Unlike using IP watermark to identify IP＇s ownership, IP fingerprinting can be used to trace illegal distributor. In this paper, IP buyer＇s fingerprint is mapped into different derived instances of extracted modules, and then is embedded into IP to identify distributor in case of illegal distribution. Comparing with other fingerprinting method, the proposed method has some good characteristics such as low design effort, small storage demand, high security and few physical overheads.     

Protein phosphatase 2A protects centromeric sister chromatid cohesion during meiosis I

Segregation of homologous maternal and paternal centromeres to opposite poles during meiosis I depends on post-replicative crossing over between homologous non-sister chromatids, which creates chiasmata and therefore bivalent chromosomes. Destruction of sister chromatid cohesion along chromosome arms due to proteolytic cleavage of cohesin's Rec8 subunit by separase resolves chiasmata and thereby triggers the first meiotic division. This produces univalent chromosomes, the chromatids of which are held together by centromeric cohesin that has been protected from separase by shugoshin (Sgo1/MEI-S332) proteins. Here we show in both fission and budding yeast that Sgo1 recruits to centromeres a specific form of protein phosphatase 2A (PP2A). Its inactivation causes loss of centromeric cohesin at anaphase I and random segregation of sister centromeres at the second meiotic division. Artificial recruitment of PP2A to chromosome arms prevents Rec8 phosphorylation and hinders resolution of chiasmata. Our data are consistent with the notion that efficient cleavage of Rec8 requires phosphorylation of cohesin and that this is blocked by PP2A at meiosis I centromeres.     

A phosphatase complex that dephosphorylates gammaH2AX regulates DNA damage checkpoint recovery

One of the earliest marks of a double-strand break (DSB) in eukaryotes is serine phosphorylation of the histone variant H2AX at the carboxy-terminal SQE motif to create gammaH2AX-containing nucleosomes. Budding-yeast histone H2A is phosphorylated in a similar manner by the checkpoint kinases Tel1 and Mec1 (ref. 2; orthologous to mammalian ATM and ATR, respectively) over a 50-kilobase region surrounding the DSB. This modification is important for recruiting numerous DSB-recognition and repair factors to the break site, including DNA damage checkpoint proteins, chromatin remodellers and cohesins. Multiple mechanisms for eliminating gammaH2AX as DNA repair completes are possible, including removal by histone exchange followed potentially by degradation, or, alternatively, dephosphorylation. Here we describe a three-protein complex (HTP-C, for histone H2A phosphatase complex) containing the phosphatase Pph3 that regulates the phosphorylation status of gammaH2AX in vivo and efficiently dephosphorylates gammaH2AX in vitro. gammaH2AX is lost from chromatin surrounding a DSB independently of the HTP-C, indicating that the phosphatase targets gammaH2AX after its displacement from DNA. The dephosphorylation of gammaH2AX by the HTP-C is necessary for efficient recovery from the DNA damage checkpoint.     

一种蛋白A的Z结构域衍生物的构建及对IgG亲和力的研究

蛋白A是金葡菌中一种重要的致病因子,有较强结合多种哺乳动物的IgG的能力,因此降低机体产生特异性SPA抗体的水平,使金葡菌具免疫逃逸功能.研究一种与IgG亲和力较弱的蛋白A衍生物,对金葡菌疫苗的开发具重要意义.研究表明,蛋白A的Z结构域中位于第13、14位的Phe,Tyr是两个与IgG结合的关键位点.本文首次通过PCR突变技术,将这两个关键氨基酸均改造为Gly,构建突变体ZFY.Discovery Studio3.5受体-配体相互作用力模块分析显示,突变体ZFY与IgG结合力降低到-61.68kcal/mol,仅为原相互作用力的10.69%;构建得到ZFY蛋白,经ITC测定,ZFY与兔IgG的亲和常数降低到1.2×104 M-1,仅为原亲和常数的0.39%,证明了ZFY是一种与IgG亲和力较弱的蛋白A衍生物,为实现新型蛋白A疫苗治疗金葡菌感染奠定基础.     

Cell cycle oscillation of phosphatase inhibitor-2 in rat fibroblasts coincident with p34cdc2 restriction

Attention has focused on the regulation of the eucaryotic cell division cycle since the protein kinase p34cdc2 was identified as a key enzyme in mitotic induction. The level of this kinase remains constant throughout the cell cycle but its activity alters, particularly before M phase. Although the factors regulating cdc2 activity are still unknown, there is increasing evidence that it is influenced by p34cdc2 dephosphorylation. Protein phosphatase inhibitor-2 (I2) is a specific inhibitor of phosphatase type-1, which with type-2A is one of the two principal Ser(P) and Thr(P) phosphatases. Here we show that the level of I2, assayed by immunofluorescence staining, activity measurements, western immunoblotting and metabolic labelling, oscillates during the cell cycle in rat fibroblasts, peaking at S phase and mitosis. Moreover, when we inhibited I2 in vivo by microinjection of anti-I2 antibodies in S-phase cells, the pseudo-mitotic cellular response to injected p34cdc2 was restored, indicating that I2 might have a role in the modulation of p34cdc2 activity.     

A surprising simplicity to protein folding

The polypeptide chains that make up proteins have thousands of atoms and hence millions of possible inter-atomic interactions. It might be supposed that the resulting complexity would make prediction of protein structure and protein-folding mechanisms nearly impossible. But the fundamental physics underlying folding may be much simpler than this complexity would lead us to expect folding rates and mechanisms appear to be largely determined by the topology of the native (folded) state, and new methods have shown great promise in predicting protein-folding mechanisms and the three-dimensional structures of proteins.     

A bacterial calcium-binding protein homologous to calmodulin

Many of the effects of calcium ions in eukaryotic cells are mediated by calcium-binding regulatory proteins such as calmodulin, in which each calcium-binding site has a distinctive helix-loop-helix conformation termed the EF hand. Protein S from the spore coat of the Gram-negative bacterium Myxococcus xanthus has been shown to resemble calmodulin in its internally-duplicated structure and ability to bind calcium. However, it has a beta-sheet secondary structure rather than the helix-loop-helix arrangement of the eukaryotic proteins. We have determined the complete amino-acid sequence of a calcium-binding protein from the Gram-positive bacterium "Streptomyces erythraeus" by cloning and sequencing the corresponding gene. It contains four EF-hand motifs bearing remarkable sequence similarity to the calcium-binding sites in calmodulin. This implies that the EF-hand super-family may have evolved from ancient proteins present in prokaryotes.     

一名鲁迅卫士的光辉足迹——评《许广平文集》

1998年出版的三巨册近百万字的《许广平文集》包括三部分:回忆和纪念鲁迅以及记叙作者自女师大时代就开始的与鲁迅并肩战斗、工作与生活;有关鲁迅著作出版的筹划、介绍以及拯救鲁迅藏书、评论研究鲁迅的著作等;作者冲出封建大家庭,走向社会,成长为中国妇女解放运动的先驱之一。这部内容丰富的巨著,清晰地记录了一位终生不渝勇敢捍卫鲁迅精神,保卫鲁迅文化遗产的坚强卫士的光辉足迹。     

Binding of the SARS-CoV-2 spike protein to glycans

The pandemic of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a high number of deaths in the world.To combat it,it is necessary to...