PDBsum entry 2c4b

Fusion protein

PDB id: 2c4b

Contents

Protein chains

142 a.a. *

Ligands

2PE ×4

GOL

EDO ×10

FMT ×3

SO4 ×15

UNX-UNX-UNX-UNX-UNX

UNX-UNX-UNX-UNX-UNX-UNX-UNX-UNX-UNX

UNX

MES

SO4-SO4

Waters ×509

* Residue conservation analysis

PDB id:

2c4b

Name:

Fusion protein

Title:

Inhibitor cystine knot protein mcoeeti fused to the catalytically inactive barnase mutant h102a

Structure:

Barnase mcoeeti fusion. Chain: a, b. Engineered: yes. Mutation: yes. Other_details: mcoeeti is a hybrid of mcoti-ii and eeti-ii. It was fused c-terminally to the h102a mutant of barnase with a 4-residue linker

Source:

Bacillus amyloliquefaciens, momordica cochinchinensis, ecballium elaterium. Organism_taxid: 1390, 3674, 3679. Expressed in: escherichia coli. Expression_system_taxid: 562. Other_details: mcoeeti is a designed hybrid carrying sequence derived from momordica cochinchinensis mcoti-ii and ecballium elaterium eeti-ii

Resolution:

1.30Å R-factor: 0.133 R-free: 0.164

Authors:

H.H.Niemann,H.U.Schmoldt,A.Wentzel,H.Kolmar,D.W.Heinz

Key ref:

H.H.Niemann et al. (2006). Barnase fusion as a tool to determine the crystal structure of the small disulfide-rich protein McoEeTI. J Mol Biol, 356, 1-8. PubMed id: 16337652 DOI: 10.1016/j.jmb.2005.11.005

Date:

18-Oct-05 Release date: 21-Nov-05

Protein chains

P00648  (RNBR_BACAM) -  Ribonuclease from Bacillus amyloliquefaciens

Seq: 157 a.a.

Struc: 142 a.a.*

Protein chains

P12071  (ITR2_ECBEL) -  Trypsin inhibitor 2 from Ecballium elaterium

Seq: 30 a.a.

Struc: 142 a.a.*

Protein chains

P82409  (ITR2_MOMCO) -  Trypsin inhibitor 2 from Momordica cochinchinensis

Seq: 34 a.a.

Struc: 142 a.a.*

* PDB and UniProt seqs differ at 18 residue positions (black crosses)

Enzyme reactions

• Enzyme class 2: E.C.3.1.27.- - ?????
• Enzyme class 3: E.C.3.1.27.3 - Transferred entry: 4.6.1.24.
• Reaction: Two-stage endonucleolytic cleavage to nucleoside 3'-phosphates and 3'-phosphooligonucleotides ending in G-P with 2',3'-cyclic phosphate intermediates.

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

DOI no: 10.1016/j.jmb.2005.11.005

J Mol Biol 356:1-8 (2006)

PubMed id: 16337652

Barnase fusion as a tool to determine the crystal structure of the small disulfide-rich protein McoEeTI.

H.H.Niemann, H.U.Schmoldt, A.Wentzel, H.Kolmar, D.W.Heinz.

ABSTRACT

We present a fusion system suited to determine the crystal structure of small disulfide-rich proteins. McoEeTI, a hybrid inhibitor cystine knot microprotein, was produced as a soluble fusion to a catalytically inactive variant of the RNAse barnase in Escherichia coli. Functioning as a versatile tag, barnase facilitated purification, crystallization and high-resolution structure determination. Flexibility of the linker region allows for different relative orientations of barnase and the fusion partner in two crystallographically independent molecules and may thereby facilitate crystal packing. Nevertheless, the linker region is well ordered in both molecules. This system may prove more generally useful to determine the crystal structure of peptides and small proteins.

Selected figure(s)

Figure 1.
Figure 1. Structure of the barnase-McoEeTI fusion protein. Overlay of the two crystallographically independent molecules. Chain A, barnase in blue and McoEeTI in cyan; chain B, barnase in red and McoEeTI in orange. The four residue linker is in light green or yellow. The overlay was performed on barnase, highlighting the rigid body rotation of the fusion partners relative to one another. The view is straight down the axis of rotation (shown as boxed cross) as defined by DYNDOM.13

Figure 2.
Figure 2. The linker between barnase and McoEeTI is well ordered. (a) Water molecules with very low B-factors bridge barnase, the linker and McoEeTI. Residues from barnase (Gln15 to His18) are shown with carbon atoms in purple, Ser113 from the linker in orange and Val116 and Ala132 from McoEeTI with carbon atoms in cyan. Hydrogen bonds from water molecules 1, 2, 3 and 5 are shown as broken green lines. (b) Plot of the main chain B-value by residue. The linker residues 111-114 (SSSM) have low B-values. Linker residues are indicated by a black bar.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2006, 356, 1-8) copyright 2006.

Figures were selected by an automated process.