1EFQ

Q38D mutant of LEN

Summary for 1EFQ

• Entry DOI: 10.2210/pdb1efq/pdb
• Related: 1EEU 1LVE 2LVE
• Descriptor: KAPPA-4 IMMUNOGLOBULIN (LIGHT CHAIN), URANYL (VI) ION, ZINC ION, ... (4 entities in total)
• Functional Keywords: human kappa-4 immunoglobulin light chain, mutant, monomer, uranyl ion in crystal contact, aspartic acid in beta-sheet, protein stability, immune system
• Biological source: Homo sapiens (human)
• Total number of polymer chains: 1
• Total formula weight: 12972.41

Authors

Pokkuluri, P.R.,Cai, X.,Gu, M.,Stevens, F.J.,Schiffer, M. (deposition date: 2000-02-09, release date: 2001-02-09, Last modification date: 2024-10-30)

Primary citation

Pokkuluri, P.R.,Gu, M.,Cai, X.,Raffen, R.,Stevens, F.J.,Schiffer, M.
Factors contributing to decreased protein stability when aspartic acid residues are in beta-sheet regions.
Protein Sci., 11:1687-1694, 2002

PubMed Abstract: Asp residues are significantly under represented in beta-sheet regions of proteins, especially in the middle of beta-strands, as found by a number of studies using statistical, modeling, or experimental methods. To further understand the reasons for this under representation of Asp, we prepared and analyzed mutants of a beta-domain. Two Gln residues of the immunoglobulin light-chain variable domain (V(L)) of protein Len were replaced with Asp, and then the effects of these changes on protein stability and protein structure were studied. The replacement of Q38D, located at the end of a beta-strand, and that of Q89D, located in the middle of a beta-strand, reduced the stability of the parent immunoglobulin V(L) domain by 2.0 kcal/mol and 5.3 kcal/mol, respectively. Because the Q89D mutant of the wild-type V(L)-Len domain was too unstable to be expressed as a soluble protein, we prepared the Q89D mutant in a triple mutant background, V(L)-Len M4L/Y27dD/T94H, which was 4.2 kcal/mol more stable than the wild-type V(L)-Len domain. The structures of mutants V(L)-Len Q38D and V(L)-Len Q89D/M4L/Y27dD/T94H were determined by X-ray diffraction at 1.6 A resolution. We found no major perturbances in the structures of these Q-->D mutant proteins relative to structures of the parent proteins. The observed stability changes have to be accounted for by cumulative effects of the following several factors: (1) by changes in main-chain dihedral angles and in side-chain rotomers, (2) by close contacts between some atoms, and, most significantly, (3) by the unfavorable electrostatic interactions between the Asp side chain and the carbonyls of the main chain. We show that the Asn side chain, which is of similar size but neutral, is less destabilizing. The detrimental effect of Asp within a beta-sheet of an immunoglobulin-type domain can have very serious consequences. A somatic mutation of a beta-strand residue to Asp could prevent the expression of the domain both in vitro and in vivo, or it could contribute to the pathogenic potential of the protein in vivo.

PubMed: 12070321
DOI: 10.1110/ps.4920102

Experimental method

X-RAY DIFFRACTION (1.6 Å)