Mutagenesis scan

Selected Methods Error-prone PCR Mutator strains Chemical mutagens Combinatorial cassette mutagenesis Recursive ensemble mutagenesis Scanning saturation mutagenesis... 

This new concept, first described in by Willem Siemmer in 1994 (74,75), is based on the premise that evolution works more efficiently with blocks of a sequence rather than with point mutations. Thus, rather than evolution based on a series of point mutations, as one would obtain using emor-pione polymerase chain reaction (PCR) or utlier random mutagenesis scanning procedures (76), directed evolution is based on the in vitro shuffling and reassembly of DN A sequences (74,75). 

Linker-Scanning Mutagenesis. Using linker-scanning mutagenesis (24) small sequences of DNA are removed and replaced with a synthetic restriction fragment or linker. This technique is commonly used in analysis of promoters and other control sequences in DNA, while preserving the spatial relationship between the sequences. 

Hebert CA, Vitangcol RV, Baker JB. Scanning mutagenesis of interleukin-8 identifies a cluster of residues required for receptor binding. J Biol Chem 1991 266 18989-94. 

Duchesnes CE, Murphy PM, Williams TJ, Pease JE. Alanine scanning mutagenesis of the chemokine receptor CCR3 reveals distinct extracellular residues involved in recognition of the eotaxin family of chemokines. Mol Immunol 2006 43(8) 1221-1231. 

Hallet, B., Sherratt, D.J. and Hayes, F. (1997) Pentapeptide scanning mutagenesis random insertion of a variable five amino acid cassette in a target protein. Nucleic Acids Research, 25, 1866-1867. 

Pons, J. Rajpal, A. Kirsch, J.F., Energetic analysis of an antibody/antigen interface alanine scanning mutagenesis and double mutant cycles on the HyHEL-10/lysozyme interaction, Prot. Sci. 1999, 8, 958-968. 

Barker, E. L., Perlman, M. A., Adkins, E. M Houlihan, W. J., Pristupa, Z. B Niznik, H. B and Blakely, R. D. (1998) High affinity recognition of serotonin transporter antagonists defined by species-scanning mutagenesis. An aromatic residue in transmembrane domain I dictates species-selective recognition of citalopram and mazindol. J. Biol. Chem. 273, 19459-19468. 

Mortensen, O. V., Kristensen, A. S and Wiborg, O. (2001) Species-scanning mutagenesis of the serotonin transporter reveals residues essential in selective, high-affinity recognition of antidepressants. J. Neurochem. 79, 237-247. 

Smicun, Y Campbell, S. D., Chen, M. A., Gu, H and Rudnick, G. (1999) The role of external loop regions in serotonin transport. Loop scanning mutagenesis of the serotonin transporter external domain../. Biol. Chem. 274, 36058-36064. 

A related fibril model for A/ o was proposed based on scanning proline mutagenesis (Williams et al, 2004) and molecular modeling (Guo et al., 2004). This model proposes that residues 15-21, 24-28, and 31-36 form 3 /-strands, with 2 intervening turns formed by residues 22-23 and 29-30 (Fig. 17G). Residues 17 and 34 are placed in close proximity, as double cysteine mutants at these positions form disulfide bonds on oxidation after fibrillization (Shivaprasad and Wetzel, 2004). Since fibrils with this triangular cross section would not be expected to show an H0-A... 

Linker-scanning mutagenesis, 12 518 Linnaeite, 7 209t Linoleate cross-linking, 9 146 Linoleic acid (LA), 10 828 17 664 boiling point, 5 53t... 

Loo, T.W. and Clarke, D.M. (1999) Determining the structure and mechanism of the human multidrug resistance P-glycoprotein using cysteine-scanning mutagenesis and thiol-modification techniques. Biochimica et Biophysica Acta, 1461, 315-325. 

Mingarro I, Whitley P, Lemmon MA, Von Heijne G. 1996. Ala-insertion scanning mutagenesis of the glycophorin A transmembrane helix a rapid way to map helix—helix interactions. Protein Science 5 1339. 

Cysteine Scanning Mutagenesis Mapping Binding Sites of Ligand-Gated Ion Channels... 

Cysteine scanning mutagenesis mapping binding sites of ligand-gated ion channels... 

Alternative approaches to mapping secondary structure have also been reported. These include alanine-and lysine-scanning mutagenesis, both of which do not rely on chemical modification (Yan et al., 1999 Sine et al., 2002). 

Sine SM, Wang HL, Bren NH. 2002. Lysine scanning mutagenesis delinates structural model of the nicotinic receptor ligand binding domain. J Biol Chem 277 29210-292... 

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