Crystal structure of recombinant barley cysteine endoprotease B isoform 2 (EP-B2) in complex with leupeptin

Heterologous Expression, Purification, Refolding, and Structural-Functional Characterization of EP-B2, a Self-Activating Barley Cysteine Endoprotease.

Cystein proteinase EP-B2 domain, residues 133-356

VSDLPPSVDWRQKGAVTGVKDQGKCGSCWAFSTVVSVEGINAIRTGSLVSLSEQELIDCDTADNDGCQGGLMDNAFEYIK
NNGGLITEAAYPYRAARGTCNVARAAQNSPVVVHIDGHQDVPANSEEDLARAVANQPVSVAVEASGKAFMFYSEGVFTGE
CGTELDHGVAVVGYGVAEDGKAYWTVKNSWGPSWGEQGYIRVEKDSGASGGLCGIAMEASYPVKTYSKPKPTPRRALGAR
ESLNSSSVDKLAAALEHHHHHH

1.4 M LiSO4
0.1 M HEPES, pH 7.0, VAPOR DIFFUSION, HANGING DROP, temperature 298K

Actinomycetes Streptomyces roseus MA 839-A1

THE LEUPEPTIN IS COVALENTLY CONNECTED TO ACTIVE_SITE CYS 28 OF THE ENZYME TO FORM A HEMITHIOACETAL.
THERE IS A DISCREPANCY IN THE NORINE AND PDB NUMBERING, AS NORINE COUNTS ACE AND LEU TOGETHER AS ONE RESIDUE. SO THE DBREF WILL REPORT 4 PDB RESIDUES MATCHING NORINE 3 RESIDUE

Cysteine proteinase EP-B 2 precursor (E.C.3.4.22.-), ACE-LEU-LEU-argininal (leupeptin)

EP-B2, EPB2, EPB, cysteine endoprotease, endopeptidase, leupeptin, HYDROLASE, HYDROLASE-HYDROLASE INHIBITOR COMPLEX

VSDLPPSVDWRQKGAVTGVKDQGKCGSCWAFSTVVSVEGINAIRTGSLVSLSEQELIDCDTADNDGCQGGLMDNAFEYIK
NNGGLITEAAYPYRAARGTCNVARAAQNSPVVVHIDGHQDVPANSEEDLARAVANQPVSVAVEASGKAFMFYSEGVFTGE
CGTELDHGVAVVGYGVAEDGKAYWTVKNSWGPSWGEQGYIRVEKDSGASGGLCGIAMEASYPVKTYSKPKPTPRRALGAR
ESL

BINDING SITE FOR CHAIN E OF ACE-LEU-LEU-ARGININAL (LEUPEPTIN)

BINDING SITE FOR CHAIN F OF ACE-LEU-LEU-ARGININAL (LEUPEPTIN)

BINDING SITE FOR CHAIN G OF ACE-LEU-LEU-ARGININAL (LEUPEPTIN)

BINDING SITE FOR CHAIN H OF ACE-LEU-LEU-ARGININAL (LEUPEPTIN)