2E0M

Mutant Human Ribonuclease 1 (T24L, Q28L, R31L, R32L)

Summary for 2E0M

• Entry DOI: 10.2210/pdb2e0m/pdb
• Related: 2E0J 2E0L 2E0O
• Descriptor: Ribonuclease, CADMIUM ION, CHLORIDE ION, ... (4 entities in total)
• Functional Keywords: mutant human pancreatic ribonuclease 1, hydrolase
• Biological source: Homo sapiens (human)
• Cellular location: Secreted: P07998
• Total number of polymer chains: 2
• Total formula weight: 30564.49

Authors

Yamada, H.,Tamada, T.,Kosaka, M.,Kuroki, R. (deposition date: 2006-10-10, release date: 2007-08-28, Last modification date: 2024-10-30)

Primary citation

Yamada, H.,Tamada, T.,Kosaka, M.,Miyata, K.,Fujiki, S.,Tano, M.,Moriya, M.,Yamanishi, M.,Honjo, E.,Tada, H.,Ino, T.,Yamaguchi, H.,Futami, J.,Seno, M.,Nomoto, T.,Hirata, T.,Yoshimura, M.,Kuroki, R.
'Crystal lattice engineering,' an approach to engineer protein crystal contacts by creating intermolecular symmetry: crystallization and structure determination of a mutant human RNase 1 with a hydrophobic interface of leucines
Protein Sci., 16:1389-1397, 2007

PubMed Abstract: A protein crystal lattice consists of surface contact regions, where the interactions of specific groups play a key role in stabilizing the regular arrangement of the protein molecules. In an attempt to control protein incorporation in a crystal lattice, a leucine zipper-like hydrophobic interface (comprising four leucine residues) was introduced into a helical region (helix 2) of the human pancreatic ribonuclease 1 (RNase 1) that was predicted to form a suitable crystallization interface. Although crystallization of wild-type RNase 1 has not yet been reported, the RNase 1 mutant having four leucines (4L-RNase 1) was successfully crystallized under several different conditions. The crystal structures were subsequently determined by X-ray crystallography by molecular replacement using the structure of bovine RNase A. The overall structure of 4L-RNase 1 is quite similar to that of the bovine RNase A, and the introduced leucine residues formed the designed crystal interface. To characterize the role of the introduced leucine residues in crystallization of RNase 1 further, the number of leucines was reduced to three or two (3L- and 2L-RNase 1, respectively). Both mutants crystallized and a similar hydrophobic interface as in 4L-RNase 1 was observed. A related approach to engineer crystal contacts at helix 3 of RNase 1 (N4L-RNase 1) was also evaluated. N4L-RNase 1 also successfully crystallized and formed the expected hydrophobic packing interface. These results suggest that appropriate introduction of a leucine zipper-like hydrophobic interface can promote intermolecular symmetry for more efficient protein crystallization in crystal lattice engineering efforts.

PubMed: 17586772
DOI: 10.1110/ps.072851407

Experimental method

X-RAY DIFFRACTION (1.7 Å)