Protein Engineering Techniques

This brief provides a broad overview of protein-engineering research, offering a glimpse of the most common experimental methods. It also presents various computational programs with applications that are widely used in directed evolution, computational and de novo protein design. Further, it sheds light on the advantages and pitfalls of existing methodologies and future perspectives of protein engineering techniques.

Protein engineering is a fascinating mixture of molecular biology, protein structure analysis, computation, and biochemistry, with the goal of developing useful or valuable proteins. Protein Engineering Protocols will consider the two general, but not mutually exclusive, strategies for protein engineering. The first is known as rational design, in which the scientist uses detailed knowledge of the structure and function of the protein to make desired changes. The s- ond strategy is known as directed evolution. In this case, random mutagenesis is applied to a protein, and selection or screening is used to pick out variants that have the desired qualities. By several rounds of mutation and selection, this method mimics natural evolution. An additional technique known as DNA shuffling mixes and matches pieces of successful variants to produce better results. This process mimics recombination that occurs naturally during sexual reproduction. The first section of Protein Engineering Protocols describes rational p- tein design strategies, including computational methods, the use of non-natural amino acids to expand the biological alphabet, as well as impressive examples for the generation of proteins with novel characteristics. Although procedures for the introduction of mutations have become routine, predicting and und- standing the effects of these mutations can be very challenging and requires profound knowledge of the system as well as protein structures in general.

This textbook introduces readers in an accessible and engaging way to the nuts and bolts of protein expression and engineering. Various case studies illustrate each step from the early sequence searches in online databases over plasmid design and molecular cloning techniques to protein purification and characterization. Furthermore, readers are provided with practical tips to successfully pursue a career as a protein engineer. With protein engineering being a fundamental technique in almost all molecular biology labs, the book targets advanced undergraduates and graduate students working in molecular biology, biotechnology and related scientific fields.

The design and production of novel peptides and proteins occupy pivotal positions in science and technology and will continue to do so in the 21st century. Protein Engineering and Design outlines the rapid advances in computer-based modeling, protein engineering, and methods needed for protein and peptide preparation and characterization. This indispensable reference lays the groundwork for understanding this multidisciplinary activity while providing an introduction for researchers and students to the field of protein design. Introduces and defines the techniques involved in protein engineering and design Provides a concise overview of key technologies involved and demonstrates their contributions to the specialized design and production of novel proteins and peptides

This volume details basic and advanced protocols for both stages of protein engineering: the library design phase and the identification of improved variants by screening and selection. Chapters focus on enzyme engineering using rational and semi-rational approaches. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Protein Engineering: Methods and Protocols aims to aid scientists in the planning and performance of their experiments. The chapter 'Functional Analysis of Membrane Proteins Produced by Cell-Free Translation' is open access under a CC BY 4.0 license via link.springer.com.

Site-specific mutagenesis of DNA, developed some thirty years ago, has proven to be one of the most important advances in biology. By allowing the site-specific replacement of any amino acid in a protein with one of the other nineteen amino acids, it ushered in the new era of "Protein Engineering". The field of protein engineering has, however, evolved rapidly since then and the last fifteen years have witnessed remarkable advances through the use of new chemical, biochemical and molecular biological tools towards the synthesis and manipulation of proteins. The chapters included in this book reflect the rapid evolution of protein engineering and its many applications in basic research, biotechnology, material sciences and therapy. This book will provide the reader with an introduction to state-of the-art concepts and methods and will be of use to anyone interested in the study of proteins, in academia as well as in industry.