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Creative Biostructure has developed protein evolution and engineering platform through years of experience, our seasoned scientists are specilized in providing tailored mutagenesis library construction services based on scanning point mutation technique.
Scanning point mutation is an excellent and systematic means of improving protein properties, which can replace each amino acid of target protein with all other 19 amino acids simultaneously. This powerful technique can offer a detailed profile of each amino acid at the position. For each desired codon, a small, site-saturation library can be constructed. Scanning point mutation library can be delivered by Creative Biostructure as a pool or in a separated format for any substitution variant. Creative Biostructure provides superb sequential permutation scanning library construction services.
Notably, Creative Biostructure has strong expertise in custom Alanine (Ala) Scanning serivces for protein engineering. Ala scanning technique is a popularly-used high-throughput mutagenesis method in which amino acid residues in a targe protein are systematically replaced by alanines at selected positions, including site-directed mutagenesis, expressed, and analyzed for protein function. It is reported that substitution with alanine residues can eliminate side-chain (R = methyl) interactions without changing main-chain conformation or introducing electrostatic or steric effects, therefore, which is often the preferred choice for determining the contribution of specific side-chains while retaining native and entire protein structure. Alanine scanning can be used for testing activation and coupling, interactions with ligands and monoclonal antibodies (mAbs).
Figure 1. Alanine scanning used in phage display.
Many structurally complex proteins are thousands of amino acids amino acids long, individual alanine substitutions across their entire length will be laborious and time-consuming, typically limiting functional analysis of target proteins. Thus, Creative Biostructure can use combinatorial libraries of mutant proteins to rapidly analyze protein function and identify important sidechain functionalities. Combinatorial libraries of alanine substitutions are an alternative to the laborious method of scanning individual positions in a protein. There are two approaches to perform combinatorial libraries: binomial mutagenesis and shotgun scanning. Shotgun scanning is a simplified method to identify epitope and functional mapping. Creative Biostructure ensures that using shotgun mutagenesis to engineer protein variants for improving desired properties such as enhanced expression or solubility.
E. Berdougo and B. J. Doranz (2012). High-Throughput Alanine Scanning. Genetic Engineering & Biotechnology New. Vol. 32, No. 12
K. L. Morrison and G.A. Weiss (2001). Combinatorial alanine-scanning. Curr. Opin. Chem. Biol., 5: 302-307.
L. Q. Al-Mawsawi, et al. (2014). High-throughput profiling of point mutations across the HIV-1 genome. Retrovirology, 11: 124. DOI 10.1186/s12977-014-0124-6