caa a22 4500 60549861X CHVBK 20210128100547.0 cr unu---uuuuu 210128e20151001xx s 000 0 eng 10.1007/s00253-015-6796-6 doi (NATIONALLICENCE)springer-10.1007/s00253-015-6796-6 Cysteine-free non-canonical C-intein for versatile protein C-terminal labeling through trans -splicing [Elektronische Daten] [Xudong Dai, Qijing Xun, Xiang-Qin Liu, Qing Meng] Site-specific protein labeling are powerful means of protein research and engineering; however, new and improved labeling methods are greatly needed. Split inteins catalyze a protein trans-splicing reaction that can be used for enzymatic and nearly seamless protein labeling. Non-canonical S11 split intein has been used in an earlier method of protein C-terminal labeling; however, its relatively large (~150 aa) N-intein fused to the target protein often hindered protein expression, folding, and solubility. To solve this problem, here, we have designed and demonstrated a new method of protein C-terminal labeling, by first engineering a functional non-canonical S1 split intein that has an extremely small (12 aa) N-intein and a cysteine-free C-intein. An engineered Rma DnaB S1 split intein was modified to have a cysteine-free C-intein, while still retaining its robust trans-splicing function, which permitted the C-extein in a C-precursor to have a single cysteine for easy and specific linkage with desired labeling groups. The resulting new and generally useful method has two unique advantages: (1) The extremely small (12 aa) N-intein, which must be fused to the C terminus of the target protein, is less likely to hinder the protein expression, folding, and solubility; and (2) the single cysteine in the C-extein may be readily linked to a variety of labeling or modification groups using commercially available reagents. Springer-Verlag Berlin Heidelberg, 2015 Protein labeling nationallicence Split intein nationallicence trans -splicing nationallicence Site-specific modifications nationallicence Dai Xudong College of Textiles, Donghua University, 201620, Shanghai, China aut Xun Qijing College of Textiles, Donghua University, 201620, Shanghai, China aut Liu Xiang-Qin Department of Biochemistry & Molecular Biology, Dalhousie University, B3H 4R2, Halifax, Nova Scotia, Canada aut Meng Qing College of Textiles, Donghua University, 201620, Shanghai, China aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/19(2015-10-01), 8151-8161 0175-7598 99:19<8151 2015 99 253 https://doi.org/10.1007/s00253-015-6796-6 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6796-6 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Dai Xudong College of Textiles, Donghua University, 201620, Shanghai, China aut NATIONALLICENCE 700 1- Xun Qijing College of Textiles, Donghua University, 201620, Shanghai, China aut NATIONALLICENCE 700 1- Liu Xiang-Qin Department of Biochemistry & Molecular Biology, Dalhousie University, B3H 4R2, Halifax, Nova Scotia, Canada aut NATIONALLICENCE 700 1- Meng Qing College of Textiles, Donghua University, 201620, Shanghai, China aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/19(2015-10-01), 8151-8161 0175-7598 99:19<8151 2015 99 253