4DWX

Crystal Structure of a Family GH-19 Chitinase from rye seeds

Summary for 4DWX

• Entry DOI: 10.2210/pdb4dwx/pdb
• Related: 4DYG
• Descriptor: Basic endochitinase C, 2-(N-MORPHOLINO)-ETHANESULFONIC ACID, SULFATE ION, ... (5 entities in total)
• Functional Keywords: chitinase, hydrolase, carbohydrate
• Biological source: Secale cereale (Rye)
• Total number of polymer chains: 2
• Total formula weight: 53516.29

Authors

Numata, T.,Umemoto, N.,Ohnuma, T.,Fukamizo, T. (deposition date: 2012-02-27, release date: 2012-08-15, Last modification date: 2024-10-30)

Primary citation

Ohnuma, T.,Numata, T.,Osawa, T.,Inanaga, H.,Okazaki, Y.,Shinya, S.,Kondo, K.,Fukuda, T.,Fukamizo, T.
Crystal structure and chitin oligosaccharide-binding mode of a 'loopful' family GH19 chitinase from rye, Secale cereale, seeds
Febs J., 279:3639-3651, 2012

PubMed Abstract: The substrate-binding mode of a 26-kDa GH19 chitinase from rye, Secale cereale, seeds (RSC-c) was investigated by crystallography, site-directed mutagenesis and NMR spectroscopy. The crystal structure of RSC-c in a complex with an N-acetylglucosamine tetramer, (GlcNAc)(4) , was successfully solved, and revealed the binding mode of the tetramer to be an aglycon-binding site, subsites +1, +2, +3, and +4. These are the first crystallographic data showing the oligosaccharide-binding mode of a family GH19 chitinase. From HPLC analysis of the enzymatic reaction products, mutation of Trp72 to alanine was found to affect the product distribution obtained from the substrate, p-nitrophenyl penta-N-acetyl-β-chitopentaoside. Mutational experiments confirmed the crystallographic finding that the Trp72 side chain interacts with the +4 moiety of the bound substrate. To further confirm the crystallographic data, binding experiments were also conducted in solution using NMR spectroscopy. Several signals in the (1) H-(15) N HSQC spectrum of the stable isotope-labeled RSC-c were affected upon addition of (GlcNAc)(4) . Signal assignments revealed that most signals responsive to the addition of (GlcNAc)(4) are derived from amino acids located at the surface of the aglycon-binding site. The binding mode deduced from NMR binding experiments in solution was consistent with that from the crystal structure.

PubMed: 22831795
DOI: 10.1111/j.1742-4658.2012.08723.x

Experimental method

X-RAY DIFFRACTION (1.8 Å)