Multiple crystal forms ofN,N′-diacetylchitobiose deacetylase fromPyrococcus furiosus

Abstract

Native N,N 0 -diacetylchitobiose deacetylase from Pyrococcus furiosus (Pf-Dac) and its selenomethionine derivative (Se-Pf-Dac) were crystallized and analyzed in the presence and absence of cadmium ion. The four crystal structures fell into three different crystal-packing groups, with the cadmium-free Pf-Dac and Se-Pf-Dac belonging to the same space group, with homologous unit-cell parameters. The crystal structures in the presence of cadmium contained distorted octahedral cadmium complexes coordinated by three chlorides, two O atoms and an S or Se atom from the N-terminal methionine or selenomethionine, respectively. The N-terminal cadmium complex was involved in crystal contacts between symmetry-related molecules through hydrogen bonding to the N-termini. While all six N-termini of Se-Pf-Dac were involved in cadmiumcomplex formation, only two of the Pf-Dac N-termini participated in complex formation in the Cd-containing crystal, resulting in different crystal forms. These differences are discussed in light of the higher stability of the Cd-Se bond than the Cd-S bond. This work provides an example of the contribution of cadmium towards determining protein crystal quality and packing depending on the use of the native protein or the selenomethionine derivative.

Structural Insights into the Molecular Evolution of the Archaeal Exo-β-d-Glucosaminidase

“…These enzymes are thermostable, with an optimal temperature of ~80 °C, which is an important requisite for industrial applications since most industrial processes are conducted under harsh conditions (e.g., high temperature and pressure). Previous determination of the chemical structures of ChiA and Dac provided insights into their catalytic mechanism and adaptation to extremely high temperatures [4,5,6,7,8,9,10]. However, for almost 14 years after the first description of GlmA, its structure has remained unknown.…”

Secretory Expression Fine-Tuning and Directed Evolution of Diacetylchitobiose Deacetylase by Bacillus subtilis

“…To date, Dacs derived from diverse sources can be expressed in Escherichia coli to investigate the degradation mechanism of chitin (1,2,4,6); however, all the Dacs overexpressed by the plasmids were insoluble and accumulated in inclusion bodies. The gene encoding Dac was inserted into E. coli's chromosome to produce recombinant protein which is soluble in a suitable buffer solution, while the yield of Dac is extremely low and the amount of soluble protein was lower than 40 mg/liter (7).…”