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.

Design and Computational Strategies for Protein Engineering.- Combinatorial Protein Design Strategies Using Computational Methods.- Global Incorporation of Unnatural Amino Acids in Escherichia coli.- Considerations in the Design and Optimization of Coiled Coil Structures.- Calcium Indicators Based on Calmodulin-Fluorescent Protein Fusions.- Design and Synthesis of Artificial Zinc Finger Proteins.- Monobodies.- Engineering Site-Specific Endonucleases.- Evolutionary Strategies for Protein Engineering.- Protein Library Design and Screening.- Protein Design by Binary Patterning of Polar and Nonpolar Amino Acids.- Versatile DNA Fragmentation and Directed Evolution With Nucleotide Exchange and Excision Technology.- Degenerate Oligonucleotide Gene Shuffling.- M13 Bacteriophage Coat Proteins Engineered for Improved Phage Display.- Ribosome-Inactivation Display System.- Compartmentalized Self-Replication.- Synthesis of Degenerated Libraries of the Ras-Binding Domain of Raf and Rapid Selection of Fast-Folding and Stable Clones With the Dihydrofolate Reductase Protein Fragment Complementation Assay.- A General Method of Terminal Truncation, Evolution, and Re-Elongation to Generate Enzymes of Enhanced Stability.