title: Red fluorescent protein variants with incorporated non-natural amino acid analogues.
abstract: Fluorescent proteins are important tools in biotechnology applications and biosensing. DsRed, a red fluorescent protein, has expanded the colors of fluorescent proteins beyond the more commonly used green fluorescent protein. Many genetic modifications have been performed on DsRed to overcome some of its drawbacks. These primarily focused on overcoming the oligomerization detrimental to DsRed activity, and the parasitic green fluorescence caused by the immature chromophore. One such variant, DsRed-monomer, has minimal green fluorescence and no oligomerization. A few traditional mutagenesis studies have been done with DsRed and its mutants to shift the fluorescence wavelengths creating additions to the pallet of fluorescent protein colors. We have explored incorporation of non-natural amino acid analogues into DsRed-Monomer, obtaining variants with differing emission properties. In this work, two such analogues of tyrosine have been incorporated into DsRed-Monomer: 3-amino-l-tyrosine and 3-fluoro-l-tyrosine. Tyrosine analogues were chosen due to the role of tyrosine in the formation and structure of the protein's chromophore. The variants obtained in our study showed altered emission wavelengths and spectral characteristics. Our study demonstrates that incorporation of non-natural analogues into DsRed-Monomer is a viable approach to alter the spectral characteristics of the protein. We envision that this study will open up the door to non-natural mutagenesis studies with red fluorescent proteins and its mutants.
authors.affiliation: Department of Chemistry and Chemical Biology, Indiana University Purdue University, 402 N Blackford Street, LD326, Indianapolis, IN 46202, USA.
authors.forename: Ann
authors.initials: A
authors.lastname: Goulding
authors.forename: Suresh
authors.initials: S
authors.lastname: Shrestha
authors.forename: Karl
authors.initials: K
authors.lastname: Dria
authors.forename: Eric
authors.initials: E
authors.lastname: Hunt
authors.forename: Sapna K
authors.initials: SK
authors.lastname: Deo
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D000596:Amino Acids / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D008164:Luminescent Proteins / Q000737:chemistry* / Q000235:genetics* / Q000502:physiology; D015202:Protein Engineering; D013050:Spectrometry, Fluorescence; D000097573:Red Fluorescent Protein
chemical_list: D000596:Amino Acids; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein; C413663:fluorescent protein 583