Minimal domain of bacterial phytochrome required for chromophore binding and fluorescence
| Autor: | Alexander Emelyanov, Vladislav V. Verkhusha, Natalia I. Zakharova, K. A. Rumyantsev, Konstantin K. Turoverov, Daria M. Shcherbakova |
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| Přispěvatelé: | Medicum, Department of Biochemistry and Developmental Biology |
| Jazyk: | angličtina |
| Rok vydání: | 2015 |
| Předmět: |
0301 basic medicine
Molecular Sequence Data Biology behavioral disciplines and activities Article 03 medical and health sciences chemistry.chemical_compound Mice Protein structure Phycocyanobilin Bacterial Proteins Phycobilins Botany mental disorders Animals Humans Luciferase Amino Acid Sequence PLANT PHYTOCHROMES GAF DOMAIN BACTERIOPHYTOCHROME Multidisciplinary Biliverdin Spectroscopy Near-Infrared Phytochrome KNOT Biliverdine Rational design Phycocyanin Chromophore EVOLUTION PROTEIN INTERACTIONS Protein Structure Tertiary Luminescent Proteins Rhodopseudomonas 030104 developmental biology Spectrometry Fluorescence chemistry Microscopy Fluorescence Mutagenesis Biophysics 3111 Biomedicine Cyanobacteriochrome Sequence Alignment CYANOBACTERIOCHROME HeLa Cells |
| Zdroj: | Scientific Reports |
| ISSN: | 2045-2322 |
| Popis: | Fluorescent proteins (FP) are used to study various biological processes. Recently, a series of near-infrared (NIR) FPs based on bacterial phytochromes was developed. Finding ways to improve NIR FPs is becoming progressively important. By applying rational design and molecular evolution we have engineered R. palustris bacterial phytochrome into a single-domain NIR FP of 19.6 kDa, termed GAF-FP, which is 2-fold and 1.4-fold smaller than bacterial phytochrome-based NIR FPs and GFP-like proteins, respectively. Engineering of GAF-FP involved a substitution of 15% of its amino acids and a deletion of the knot structure. GAF-FP covalently binds two tetrapyrrole chromophores, biliverdin (BV) and phycocyanobilin (PCB). With the BV chromophore GAF-FP absorbs at 635 nm and fluoresces at 670 nm. With the PCB chromophore GAF-FP becomes blue-shifted and absorbs at 625 nm and fluoresces at 657 nm. The GAF-FP structure has a high tolerance to small peptide insertions. The small size of GAF-FP and its additional absorbance band in the violet range has allowed for designing a chimeric protein with Renilla luciferase. The chimera exhibits efficient non-radiative energy transfer from luciferase to GAF-FP, resulting in NIR bioluminescence. This study opens the way for engineering of small NIR FPs and NIR luciferases from bacterial phytochromes. |
| Databáze: | OpenAIRE |
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