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Butelase-1

Host Organism

Clitoria ternatea L.

Type

PAL

Sequence

MKNPLAILFLIATVVAVVSGIRDDFLRLPSQASKFFQADDNVEGTRWAVLVAGSKGYVNYRHQADVCHAYQILKKGGLKDENIIVFMYDDIAYNESNPHPGVIINHPYGSDVYKGVPKDYVGEDINPPNFYAVLLANKSALTGTGSGKVLDSGPNDHVFIYYTDHGGAGVLGMPSKPYIAASDLNDVLKKKHASGTYKSIVFYVESCESGSMFDGLLPEDHNIYVMGASDTGESSWVTYCPLQHPSPPPEYDVCVGDLFSVAWLEDCDVHNLQTETFQQQYEVVKNKTIVALIEDGTHVVQYGDVGLSKQTLFVYMGTDPANDNNTFTDKNSLGTPRKAVSQRDADLIHYWEKYRRAPEGSSRKAEAKKQLREVMAHRMHIDNSVKHIGKLLFGIEKGHKMLNNVRPAGLPVVDDWDCFKTLIRTFETHCGSLSEYGMKHMRSFANLCNAGIRKEQMAEASAQACVSIPDNPWSSLHAGFSV

External Links

PDB ID: 6DHI
EC: 3.4.22.34
MEROPS: C13 legumain
UniProt ID: A0A060D9Z7
NCBI: -

Substrate Specificity

P1
P1'
P2'
P1''
P2''
N/D
X
L/V/I/F
X except Pro
L/V/I/F

Functions

  • Intermolecular backbone terminal ligation
  • Backbone terminal cyclization, max. kcat/Km: 1.3 x 10^6 M-1s-1
  • Specificity stringency: Prime-low, Nonprime-high
  • pH optimal: pH 6-6.5

Expression Platform:

  • E. coli
  • P. pastoris

Mutations:

Other Properties:

-

References:

  1. [1] (Sequence, Activity, Substrate specificity) Nguyen, G. K. T.; Wang, S.; Qiu, Y.; Hemu, X.; Lian, Y.; Tam, J. P. Butelase 1 is an Asx-specific ligase enabling peptide macrocyclization and synthesis. Nat. Chem. Biol. 2014, 10 (9), 732-738. DOI: 10.1038/nchembio.1586
  2. [2] (Application, Kinetics) Nguyen, G. K. T.; Kam, A.; Loo, S.; Jansson, A. E.; Pan, L. X.; Tam, J. P. Butelase 1: A versatile ligase for peptide and protein macrocyclization. J. Am. Chem. Soc. Comm. 2015, 137 (49), 15398-15401. DOI: 10.1021/jacs.5b11014
  3. [3] (Application- N-terminal modification) Nguyen, G. K.; Cao, Y.; Wang, W.; Liu, C. F.; Tam, J. P. Site-Specific N-Terminal Labeling of Peptides and Proteins using Butelase 1 and Thiodepsipeptide. Angew Chem Int Ed Engl 2015, 54 (52), 15694-15698. DOI: 10.1002/anie.201506810
  4. [4] (Application-Live cell labeling) Bi, X.; Yin, J.; Nguyen, G. K. T.; Rao, C.; Halim, N. B. A.; Hemu, X.; Tam, J. P.; Liu, C. F. Enzymatic Engineering of Live Bacterial Cell Surfaces Using Butelase 1. Angew. Chem. Int. Ed. Engl. 2017, 56 (27), 7822-7825. DOI: 10.1002/anie.201703317
  5. [5] (Structure) James, A. M.; Haywood, J.; Leroux, J.; Ignasiak, K.; Elliott, A. G.; Schmidberger, J. W.; Fisher, M. F.; Nonis, S. G.; Fenske, R.; Bond, C. S.; et al. The macrocyclizing protease butelase 1 remains autocatalytic and reveals the structural basis for ligase activity. Plant J. 2019, 98 (6), 988-999. DOI: 10.1111/tpj.14293
  6. [6] (Application-bioorthogonal synthesis) Harmand, T. J.; Bousbaine, D.; Chan, A.; Zhang, X.; Liu, D. R.; Tam, J. P.; Ploegh, H. L. One-Pot Dual Labeling of IgG 1 and Preparation of C-to-C Fusion Proteins Through a Combination of Sortase A and Butelase 1. Bioconjugate Chem. 2018, 29 (10), 3245-3249. DOI: 10.1021/acs.bioconjchem.8b00563
  7. [7] (Yeast expression) Pi, N.; Gao, M.; Cheng, X.; Liu, H.; Kuang, Z.; Yang, Z.; Yang, J.; Zhang, B.; Chen, Y.; Liu, S.; et al. Recombinant Butelase-Mediated Cyclization of the p53-Binding Domain of the Oncoprotein MdmX-Stabilized Protein Conformation as a Promising Model for Structural Investigation. Biochemistry 2019, 58 (27), 3005-3015. DOI: 10.1021/acs.biochem.9b00263
  8. [8] (Bacteria expression) Hemu, X.; Zhang, X.; Nguyen, G. K. T.; To, J.; Serra, A.; Loo, S.; Sze, S. K.; Liu, C.-F.; Tam, J. P. Characterization and application of natural and recombinant butelase-1 to improve industrial enzymes by end-to-end circularization. RSC Adv. 2021, 11 (37), 23105-23112. DOI: 10.1039/d1ra03763c
  9. [9] (Bacteria expression) Zhao, J.; Fan, R.; Jia, F.; Huang, Y.; Huang, Z.; Hou, Y.; Hu, S. Q. Enzymatic Properties of Recombinant Ligase Butelase-1 and Its Application in Cyclizing Food-Derived Angiotensin I-Converting Enzyme Inhibitory Peptides. J Agric Food Chem 2021, 69 (21), 5976-5985. DOI: 10.1021/acs.jafc.1c01755