Biological applications of protein transduction technology

The impermeable nature of the cell membrane to peptides, proteins, DNA and oligonucleotides limits the therapeutic potential of these biological agents. However, the recent discovery of short cationic peptides that cross the plasma membrane efficiently is opening up new possibilities for the intracellular delivery of such agents. These peptides are commonly referred to as protein transduction domains (PTDs) and are successfully used to transport heterologous proteins, peptides and other types of cargo into cells. Several recent reports have used the membrane transducing technology in vivo to deliver biologically active cargo into various tissues. This review discusses the structure of the most commonly used PTDs and how their ability to transduce membranes is used to regulate biological functions. It also considers future directions and the potential of this technology to move from the laboratory into the clinic.     

Protein splicing and its applications

Protein splicing elements, termed inteins, provide a fertile source for innovative biotechnology tools. First harnessed for protein purification, inteins are now used to express cytotoxic proteins, to segmentally modify or label proteins, to cyclize proteins or peptides, to study structure-activity relationships and to generate reactive polypeptide termini in expressed proteins for an expanding list of chemoselective reactions, including protein ligation.     

Protein splicing mechanisms and applications

Inteins are protein splicing elements that employ standard enzyme strategies to excise themselves from precursor proteins and ligate the surrounding sequences (exteins). The protein splicing pathway consists of four nucleophilic displacements directed by the intein plus the first C-extein residue. The intein active site(s) are formed by folding of the intein within the precursor, which brings together the splice junctions and internal intein residues that assist catalysis. Inteins with non-canonical catalytic residues splice by modified pathways. Understanding intein proteolytic cleavage and ligation activities has led to the development of many novel applications in the fields of protein engineering, enzymology, microarray production, target detection and activation of transgenes in plants. Recent advances include intein-mediated attachment of proteins to solid supports for microarray or western blot analysis, linking nucleic acids to proteins and controllable splicing, which converts inteins into molecular switches.     

Biological applications of dendrimers

In the past year, significant advances have been made in the synthesis and study of glycodendrimers and peptide dendrimers. Application of these dendrimers to the study of carbohydrate-protein and protein-protein interactions has facilitated the understanding of these processes. In addition, dendrimers show great promise as DNA- and drug-delivery systems.     

Alternative splicing and protein function

Background  

Alternative splicing is a major mechanism of generating protein diversity in higher eukaryotes. Although at least half, and probably more, of mammalian genes are alternatively spliced, it was not clear, whether the frequency of alternative splicing is the same in different functional categories. The problem is obscured by uneven coverage of genes by ESTs and a large number of artifacts in the EST data.     

Functional roles of protein splicing factors

RNA splicing is one of the fundamental processes in gene expression in eukaryotes. Splicing of pre-mRNA is catalysed by a large ribonucleoprotein complex called the spliceosome, which consists of five small nuclear RNAs and numerous protein factors. The spliceosome is a highly dynamic structure, assembled by sequential binding and release of the small nuclear RNAs and protein factors. DExD/H-box RNA helicases are required to mediate structural changes in the spliceosome at various steps in the assembly pathway and have also been implicated in the fidelity control of the splicing reaction. Other proteins also play key roles in mediating the progression of the spliceosome pathway. In this review, we discuss the functional roles of the protein factors involved in the spliceosome pathway primarily from studies in the yeast system.     

Biological applications of second harmonic imaging

Second Harmonic Generation (SHG) microscopy dates back to 1974, but effective biological use of the technique has a history of barely 10 years. It is now widely used to image collagen in many different applications, and is becoming useful for imaging myosin and some polysaccharides. A separate line on research has focussed on SHG dyes, which can provide high-speed indication of membrane potential and are now in use in neurobiology. This review looks at the progress to date in these different fields.     

MicroRNA的生物学功能及其应用

MicroRNA是一类内源性的小片段的非编码RNA调控因子,广泛的存在于各种真核细胞中,参与调节细胞生长发育、分化、凋亡并影响肿瘤的发生.随着对生物治疗的深入研究,microRNA在诸多肿瘤的诊断、治疗以及抗病毒治疗方面的重要性也日渐凸显.本文就microRNA重要的生物功能及其在生物治疗中的作用进行综述和展望.     

Alternative splicing and protein structure evolution

Alternative splicing is thought to be one of the major sources for functional diversity in higher eukaryotes. Interestingly, when mapping splicing events onto protein structures, about half of the events affect structured and even highly conserved regions i.e. are non-trivial on the structure level. This has led to the controversial hypothesis that such splice variants result in nonsense-mediated mRNA decay or non-functional, unstructured proteins, which do not contribute to the functional diversity of an organism. Here we show in a comprehensive study on alternative splicing that proteins appear to be much more tolerant to structural deletions, insertions and replacements than previously thought. We find literature evidence that such non-trivial splicing isoforms exhibit different functional properties compared to their native counterparts and allow for interesting regulatory patterns on the protein network level. We provide examples that splicing events may represent transitions between different folds in the protein sequence–structure space and explain these links by a common genetic mechanism. Taken together, those findings hint to a more prominent role of splicing in protein structure evolution and to a different view of phenotypic plasticity of protein structures.     

Biological applications of spin pH probes.

The determination of pH is one of the most important problems in the biochemistry of living organisms, since many of the vital processes of cells and cellular organelles depend on the local pH value. Amongst currently used experimental approaches for the measurement of pH, the application of spin pH probes in combination with EPR spectroscopy is a comparatively new and rapidly developing field. In this article we describe the background, advantages and limitations of the method of spin pH probes, and discuss its recent applications. Availability of a wide variety of pH-sensitive nitroxides with different ranges of pH-sensitivity, labeling group and lipophilicity facilitates their application to a variety of biological systems from subcellular organelles to complex organisms. The recent progress in low-field EPR-based imaging and spectroscopy-based techniques allows spin pH probes to be used for non-invasive in vivo pH measurement and pH-sensitive imaging.     

Biological applications of synchrotron radiation infrared spectromicroscopy

Extremely brilliant infrared (IR) beams provided by synchrotron radiation sources are now routinely used in many facilities with available commercial spectrometers coupled to IR microscopes. Using these intense non-thermal sources, a brilliance two or three order of magnitude higher than a conventional source is achievable through small pinholes (< 10 μm) with a high signal to-noise ratio. IR spectroscopy is a powerful technique to investigate biological systems and offers many new imaging opportunities. The field of infrared biological imaging covers a wide range of fundamental issues and applied researches such as cell imaging or tissue imaging. Molecular maps with a spatial resolution down to the diffraction limit may be now obtained with a synchrotron radiation IR source also on thick samples. Moreover, changes of the protein structure are detectable in an IR spectrum and cellular molecular markers can be identified and used to recognize a pathological status of a tissue. Molecular structure and functions are strongly correlated and this aspect is particularly relevant for imaging. We will show that the brilliance of synchrotron radiation IR sources may enhance the sensitivity of a molecular signal obtained from small biosamples, e.g., a single cell, containing extremely small amounts of organic matter. We will also show that SR IR sources allow to study chemical composition and to identify the distribution of organic molecules in cells at submicron resolution is possible with a high signal-to-noise ratio. Moreover, the recent availability of two-dimensional IR detectors promises to push forward imaging capabilities in the time domain. Indeed, with a high current synchrotron radiation facility and a Focal Plane Array the chemical imaging of individual cells can be obtained in a few minutes. Within this framework important results are expected in the next years using synchrotron radiation and Free Electron Laser (FEL) sources for spectro-microscopy and spectral-imaging, alone or in combination with Scanning Near-field Optical Microscopy methods to study the molecular composition and dynamic changes in samples of biomedical interest at micrometric and submicrometric scales, respectively.     

SR蛋白家族在RNA剪接中的调控作用

SR蛋白家族成员都具有一个富含丝氨酸/精氨酸(S/R)重复序列的RS结构域,在RNA剪接体的组装和选择性剪接的调控过程中具有重要的作用。绝大多数SR蛋白是生存的必需因子,通过其RS结构域和特有的其他结构域,实现与前体mRNA的特异性序列或其他剪接因子的相互作用,协同完成剪接位点的正确选择或促进剪接体的形成。深入研究SR蛋白家族在RNA选择性剪接中的调控机制,可以促进以疾病治疗或害虫防治为目的的应用研究。该文总结了SR蛋白家族在基础研究和应用方面的进展。