Enzyme engineering is a valuable tool for generating powerful and robust biocatalysts for a broad range of applications. Directed evolution using cell surface display has been previously applied for enzyme engineering, and involves iterative optimization of a protein sequence to yield a desired phenotype. However, previous applications have been carefully tailored for specific enzymes, and this process of developing a good screen is a significant bottleneck in engineering. Co-display of an enzyme and its substrate on the yeast surface is a promising general strategy for evolving bond-forming enzymes, but has only been used to engineer one enzyme. We extended yeast surface co-display to engineer Sfp, a bioconjugation enzyme, for improved activity with a non-native substrate. Sfp library screening identified mutational hotspots in the enzyme and led to the design of Sfp variants with orders of magnitude enhancements in activity. However, these activity improvements were only apparent in contexts similar to yeast surface co-display, reflecting artifacts in the use of this approach for enzyme engineering. Based on our results, we offer suggestions for improving the success of engineering strategies based on yeast surface display.

Bioconjugate Techniques, 2nd Edition, is the essential guide to the modification and cross linking of biomolecules for use in research, diagnostics, and therapeutics. It provides highly detailed information on the chemistry, reagent systems, and practical applications for creating labeled or conjugate molecules. It also describes dozens of reactions with details on hundreds of commercially available reagents and the use of these reagents for modifying or cross linking peptides and proteins, sugars and polysaccharides, nucleic acids and oligonucleotides, lipids, and synthetic polymers. A one-stop source for proven methods and protocols for synthesizing bioconjugates in the lab Step-by-step presentation makes the book an ideal source for researchers who are less familiar with the synthesis of bioconjugates More than 600 figures that visually describe the complex reactions associated with the synthesis of bioconjugates Includes entirely new chapters on the latest areas in the field of bioconjugation as follows: Microparticles and nanoparticlesSilane coupling agentsDendrimers and dendronsChemoselective ligationQuantum dotsLanthanide chelatesCyanine dyesDiscrete PEG compoundsBuckyballs,fullerenes, and carbon nanotubesMass tags and isotope tagsBioconjugation in the study of protein interactions

Enzyme Engineering An authoritative and up-to-date discussion of enzyme engineering and its applications In Enzyme Engineering: Selective Catalysts for Applications in Biotechnology, Organic Chemistry, and Life Science, a team of distinguished researchers deliver a robust treatment of enzyme engineering and its applications in various fields such as biotechnology, life science, and synthesis. The book begins with an introduction to different protein engineering techniques, covers topics like gene mutagenesis methods for directed evolution and rational enzyme design. It includes industrial case studies of enzyme engineering with a focus on selectivity and activity. The authors also discuss new and innovative areas in the field, involving machine learning and artificial intelligence. It offers several insightful perspectives on the future of this work. Readers will also find: A thorough introduction to directed evolution and rational design as protein engineering techniques Comprehensive explorations of screening and selection techniques, gene mutagenesis methods in directed evolution, and guidelines for applying gene mutagenesis in organic chemistry, pharmaceutical applications, and biotechnology Practical discussions of protein engineering of enzyme robustness relevant to organic and pharmaceutical chemistry Treatments of artificial enzymes as promiscuous catalysts Various lessons learned from semi-rational and rational directed evolution A transdisciplinary treatise, Enzyme Engineering: Selective Catalysts for Applications in Biotechnology, Organic Chemistry, and Life Science is perfect for protein engineers, theoreticians, organic, and pharmaceutical chemists as well as transition metal researchers in catalysis and biotechnologists.

Isolated enzymes are known as useful for synthesising of complex molecules with more and more large scale applications in recent years. These six reviews show different steps in the development of enzymatic syntheses for useful compounds. Screening of novel enzymes, their biochemical characterisation as well as reaction engineering approaches in order to increase their performance are treated in three papers. Then the use of protein design as a tool for the directed improvement of reactivity and selectivity of biocatalysts is described in two reviews. The supply of sufficient quantities of biocatalysts must not be neglected: fluidized bed adsorption as an emerging technology for the simple and fast purification of biomolecules is presented in the light of its potential in large scale supply of enzyme catalysts.

Isolated enzymes are known as useful for synthesising of complex molec ules with more and more large scale applications in recent years. Thes e six reviews show different steps in the development of enzymatic syn theses for useful compounds. Screening of novel enzymes, their biochem ical characterisation as well as reaction engineering approaches in or der to increase their performance are treated in three papers. Then th e use of protein design as a tool for the directed improvement of reac tivity and selectivity of biocatalysts is described in two reviews. Th e supply of sufficient quantities of biocatalysts must not be neglecte d: fluidized bed adsorption as an emerging technology for the simple a nd fast purification of biomolecules is presented in the light of its potential in large scale supply of enzyme catalysts.

Enzyme Technology is one the most promising disciplines in modern biotechnology. In this book, the applications of a wide variety of enzymes are highlighted. Current studies in enzyme technology are focused towards the discovery of novel enzymes (termed “bio-discovery” or “bio-prospecting”) and the identification and elucidation of novel pathways of these novel enzymes with emphasis on their industrial relevance. With the development of molecular techniques and other bioinformatics tools, the time to integrate this subject with other fields in the life sciences has arrived. A rapid expansion of the knowledge base in the field of enzyme biotechnology has occurred over the past few years. Much of this expansion has been driven by the bio-discovery of many new enzymes from a wide range of environments, some extreme in nature, followed by subsequent protein (enzyme) engineering. These enzymes have found a wide range of applications, ranging from bioremediation, bio-monitoring, biosensor development, bioconversion to biofuels and other biotechnologically important value-added products. Hydrolases constitute a major component of the global annual revenue generated by industrial enzymes and the emphasis has therefore been placed on these enzymes and their applications. With the immense interest of researchers active in this area, this book will serve to provide information on current aspects in this field of study. In the current edition, the contributions of many diversified topics towards establishing new directions of research in the area of enzyme biotechnology are described. This book serves to provide a unique source of information to undergraduates, post graduates and doctoral courses in microbiology and biotechnology along with allied life sciences. The present edition of the book covers all important areas of enzyme biotechnology i.e. the wide variety of enzymes in the field of enzyme biotechnology and their industrial applications, new methods and state-of-the-art information on modern methods of enzyme discovery. This book will act as good resource on most of the current facets of enzyme technology for all students engaged in bioengineering and biotechnology.

In this Completely Revised and Extended Edition with a significantly enhanced content, all Chapters have been updated considering relevant literature and recent developments until 2016 together with application oriented examples with a focus on Industrial Biocatalysis. Newly treated topics comprise among others systems metabolic engineering approaches, metagenome screening, new tools for pathway engineering, and de-novo computational design as actual research areas in biocatalysis. Information about different aspects of RNA technologies, and completely new Chapters on 'Fluorescent Proteins' and 'Biocatalysis and Nanotechnology' are also included.