PDBsum entry: 1b6v

Molecular evolution

PDB id: 1b6v

Contents

Protein chains

124 a.a. *

Waters ×102

* Residue conservation analysis

PDB id:

1b6v

Name:

Molecular evolution

Title:

Crystal structure of a hybrid between ribonuclease a and bovine seminal ribonuclease

Structure:

Ribonuclease. Chain: a, b. Engineered: yes. Mutation: yes

Source:

Bos taurus. Cattle. Organism_taxid: 9913. Cell_line: lon- hptr- tetr cells. Expressed in: escherichia coli. Expression_system_taxid: 562. Expression_system_cell_line: lon- hptr- tetr cells. Other_details: synthetic gene

Resolution:

2.00Å R-factor: 0.182 R-free: 0.256

Authors:

E.H.Vatzaki,S.C.Allen,D.D.Leonidas,K.R.Acharya

Key ref:

E.H.Vatzaki et al. (1999). Crystal structure of a hybrid between ribonuclease A and bovine seminal ribonuclease--the basic surface, at 2.0 A resolution. Eur J Biochem, 260, 176-182. PubMed id: 10091597 DOI: 10.1046/j.1432-1327.1999.00142.x

Date:

18-Jan-99 Release date: 15-Jun-99

Protein chains

P61823  (RNAS1_BOVIN) -  Ribonuclease pancreatic

Seq: 150 a.a.

Struc: 124 a.a.*

* PDB and UniProt seqs differ at 7 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.1.27.5 - Pancreatic ribonuclease.
• Reaction: Endonucleolytic cleavage to nucleoside 3'-phosphates and 3'-phosphooligonucleotides ending in C-P or U-P with 2',3'-cyclic phosphate intermediates.

 Gene Ontology (GO) functional annotation 

• Cellular component: extracellular region (1 term)
• Biochemical function: nucleic acid binding (6 terms)

DOI no: 10.1046/j.1432-1327.1999.00142.x

Eur J Biochem 260:176-182 (1999)

PubMed id: 10091597

Crystal structure of a hybrid between ribonuclease A and bovine seminal ribonuclease--the basic surface, at 2.0 A resolution.

E.H.Vatzaki, S.C.Allen, D.D.Leonidas, K.Trautwein-Fritz, J.Stackhouse, S.A.Benner, K.R.Acharya.

ABSTRACT

A variant of bovine pancreatic ribonuclease A has been prepared with seven amino acid substitutions (Q55K, N62K, A64T, Y76K, S80R, E111G, N113K). These substitutions recreate in RNase A the basic surface found in bovine seminal RNase, a homologue of pancreatic RNase that diverged some 35 million years ago. Substitution of a portion of this basic surface (positions 55, 62, 64, 111 and 113) enhances the immunosuppressive activity of the RNase variant, activity found in native seminal RNase, while substitution of another portion (positions 76 and 80) attenuates the activity. Further, introduction of Gly at position 111 has been shown to increase the catalytic activity of RNase against double-stranded RNA. The variant and the wild-type (recombinant) protein were crystallized and their structures determined to a resolution of 2.0 A. Each of the mutated amino acids is seen in the electron density map. The main change observed in the mutant structure compared with the wild-type is the region encompassing residues 16-22, where the structure is more disordered. This loop is the region where the polypeptide chain of RNase A is cleaved by subtilisin to form RNase S, and undergoes conformational change to allow residues 1-20 of the RNase to swap between subunits in the covalent seminal RNase dimer.

Selected figure(s)

Figure 1.
Fig. 1. Surface representation of wild-type RNase A (A) and the variant (B) dimers in the crystallographic asymmetric unit. The surfaces are coloured according to electrostatic potential from electronegative to electropositive by a red-to-blue continuous colour range. The views of mol I in both figures are almost identical: view in (B) shows the `basic surface' in the variant and the view in (A) shows the corresponding surface in the wild-type RNase A molecule. Figure was generated using GRASP [44] with a probe radius of 1.4 Å.

Figure 2.
Fig. 2. Stereoview of the C^ backbone for the variant RNase molecule. Side chains of the mutated residues are shown as ball-and-stick models.

The above figures are reprinted by permission from the Federation of European Biochemical Societies: Eur J Biochem (1999, 260, 176-182) copyright 1999.