4wqq

From Proteopedia

Structure of EPNH mutant of CEL-I

Structural highlights

4wqq is a 4 chain structure with sequence from Cucumaria echinata. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
Method:	X-ray diffraction, Resolution 1.7Å
Ligands:	,
Resources:	FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT

Function

Q7M462_CUCEC

Publication Abstract from PubMed

BACKGROUND: CEL-I is a galactose/N-acetylgalactosamine-specific C-type lectin isolated from the sea cucumber Cucumaria echinata. Its carbohydrate-binding site contains a QPD (Gln-Pro-Asp) motif, which is generally recognized as the galactose specificity-determining motif in the C-type lectins. In our previous study, replacement of the QPD motif by an EPN (Glu-Pro-Asn) motif led to a weak binding affinity for mannose. Therefore, we examined the effects of an additional mutation in the carbohydrate-binding site on the specificity of the lectin. METHODS: Trp105 of EPN-CEL-I was replaced by a histidine residue using site-directed mutagenesis, and the binding affinity of the resulting mutant, EPNH-CEL-I, was examined by sugar-polyamidoamine dendrimer assay, isothermal titration calorimetry, and glycoconjugate microarray analysis. Tertiary structure of the EPNH-CEL-I/mannose complex was determined by X-ray crystallographic analysis. RESULTS: Sugar-polyamidoamine dendrimer assay and glycoconjugate microarray analysis revealed a drastic change in the specificity of EPNH-CEL-I from galactose/N-acetylgalactosamine to mannose. The association constant of EPNH-CEL-I for mannose was determined to be 3.17x103M-1 at 25 degrees C. Mannose specificity of EPNH-CEL-I was achieved by stabilization of the binding of mannose in a correct orientation, in which the EPN motif can form proper hydrogen bonds with 3- and 4-hydroxy groups of the bound mannose. CONCLUSIONS: Specificity of CEL-I can be engineered by mutating a limited number of amino acid residues in addition to the QPD/EPN motifs. GENERAL SIGNIFICANCE: Versatility of the C-type carbohydrate-recognition domain structure in the recognition of various carbohydrate chains could become a promising platform to develop novel molecular recognition proteins.

Mannose-recognition mutant of the galactose/N-acetylgalactosamine-specific C-type lectin CEL-I engineered by site-directed mutagenesis.,Moriuchi H, Unno H, Goda S, Tateno H, Hirabayashi J, Hatakeyama T Biochim Biophys Acta. 2015 Apr 11;1850(7):1457-1465. doi:, 10.1016/j.bbagen.2015.04.004. PMID:25869490^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

References

↑ Moriuchi H, Unno H, Goda S, Tateno H, Hirabayashi J, Hatakeyama T. Mannose-recognition mutant of the galactose/N-acetylgalactosamine-specific C-type lectin CEL-I engineered by site-directed mutagenesis. Biochim Biophys Acta. 2015 Apr 11;1850(7):1457-1465. doi:, 10.1016/j.bbagen.2015.04.004. PMID:25869490 doi:http://dx.doi.org/10.1016/j.bbagen.2015.04.004