title: Incorporation of Non-Canonical Amino Acids.
abstract: In this chapter we discuss the strengths, caveats and technical considerations of three approaches for reprogramming the chemical composition of selected amino acids within a membrane protein. In vivo nonsense suppression in the Xenopus laevis oocyte, evolved orthogonal tRNA and aminoacyl-tRNA synthetase pairs and protein ligation for biochemical production of semisynthetic proteins have been used successfully for ion channel and receptor studies. The level of difficulty for the application of each approach ranges from trivial to technically demanding, yet all have untapped potential in their application to membrane proteins.
authors.affiliation: Department of Molecular Physiology and Biophysics, University of Iowa, 51 Newton Road, 52246, Iowa City, IA, USA.
authors.forename: Lilia
authors.initials: L
authors.lastname: Leisle
authors.affiliation: Department of Physiology and Pharmacology, Oregon Health and Sciences University, 97239, Portland, OR, USA.
authors.forename: Francis
authors.initials: F
authors.lastname: Valiyaveetil
authors.affiliation: Department of Biochemistry and Biophysics, Oregon State University Corvallis, 97331, Corvallis, OR, USA.
authors.forename: Ryan A
authors.initials: RA
authors.lastname: Mehl
authors.affiliation: Department of Molecular Physiology and Biophysics, University of Iowa, 51 Newton Road, 52246, Iowa City, IA, USA. christopher-ahern@uiowa.edu.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Ahern
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism*; D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism; D000818:Animals; D003062:Codon; D006801:Humans; D047668:Inteins; D007473:Ion Channels / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D015202:Protein Engineering / Q000379:methods*; D013329:Structure-Activity Relationship; D014982:Xenopus laevis / Q000235:genetics / Q000378:metabolism
chemical_list: D000596:Amino Acids; D003062:Codon; D007473:Ion Channels; D000604:Amino Acyl-tRNA Synthetases
keywords: Evolved tRNA synthetase pairs; Intein ligation; Nonsense suppression; Protein semi-synthesis