Protein Expression And Purification
Knowledge of the three-dimensional structure of a protein is absolutely required for the complete understanding of its function. The spatial orientation of amino acids in the active site of an enzyme demonstrates how substrate specificity is defined, and assists the medicinal chemist in the design of s- cific, tight-binding inhibitors. The shape and contour of a protein surface hints at its interaction with other proteins and with its environment.
The expression of proteins from different host organisms is of big interest in the modern biotechnology. Not only for commercial or medical purposes, but also for the characterization of foreign proteins the heterologous expression plays a significant role.How can a foreign protein be expressed in a pro- or eukaryotic host? Which host is suitable for which kind of protein? How can the specific protein yield be increased by varying process parameter?The author Dr.
Tellurite resistance (TeR) determinat from uropathogenic strain Escherichia coli KL53 is harbored on conjugative plasmid pTE53, which consists of two clusters of ter genes. The first cluster of genes is responsible for the TeR terZABCDEF and protective region consists of terXYW. Only four genes terBCDE are essential for TeR and these genes were object of our study.
A recombinant S100A4 protein was produced in Pichia pastoris (Pichia). The expression vector contained parts of the AOX1 promoter and also a secretion signal, making it possible to purify the protein from the yeast media after methanol induction. Attempts were made to purify the protein from the media both by hydrophobic interaction chromatography (HIC) and by anionic ion exchange chromatography (IEC).
On the forefront of modern scientific innovation, Cloning, Gene Expression and Protein Purification: Experimental Procedures and Process Rationale effectively doubles as a laboratory manual for students and a reference book for professional researchers. Designed for advanced undergraduate and beginning graduate students in molecular biology, this unique combination lecture/laboratory resource presents detailed protocols for the multi-step process involved in isolating a gene, cloning and characterizing it, expressing its encoded protein, and purifying and characterizing the protein's basic physical properties.
This manual is an indispensable tool for introducing advanced undergraduates and beginning graduate students to the techniques of recombinant DNA technology, or gene cloning and expression. The techniques used in basic research and biotechnology laboratories are covered in detail. Students gain hands-on experience from start to finish in subcloning a gene into an expression vector, through purification of the recombinant protein.
Despite exciting advances in genome sequencing, isolating a protein from its expression system in its native form still presents a complex challenge. In High Throughput Protein Expression and Purification: Methods and Protocols, leading scientists detail the most successful protocols currently in use, including various high throughput cloning schemes, protein expression analysis, and production protocols.
HIV Volume 2 explores the biochemistry and molecular biology of HIV. Methods for cloning and expressing most of the viral proteins are described as well as assays for studying enzyme function in vitro. It also includes a section on recent approaches to drug discovery. Contents: The HIV life cycle; Reverse transcriptase; Ribonulease H; Integrase; Protease; Expression of gag, envelope and other HIV antigens using Bacculovirus; Envelope expression and purification; HIV envelope glyocoprotein/CD4 interactions - studies using recombinant vaccinia virus vectors; RNA binding assays for the regulatory proteins tat and rev; Cellular and cell-free assays for tat; Cellular assays for rev; DNA binding proteins interacting with the viral LTR; Nef; Vpu; Vif; Cellular assays for antiviral drugs; Inhibition of HIV by ribozymes and antisense oligonucleotides; Gene therapy approaches to the inhibition of HIV