BER Structural Biology and Imaging Resources
Synchrotron, Neutron, and Cryo-EM
U.S. Department of Energy | Office of Science | Office of Biological and Environmental Research

Structural Characterization of Chitinases from Agave tequilana

July 10, 2019

Chitinases are enzymes that hydrolyze chitin, a molecule present in fungal cell walls, most insect exoskeletons, yeasts and algae, and crustacean shells, among other organisms. These enzymes are involved in numerous physiological events such as abiotic stress responses, including antifungal activity and defense mechanisms. The genus Agave is cultivated to produce fiber, food, beverages, and fuel, for example, and has great socioeconomic and agroecological value, especially in hot and drought-prone regions of the world. Despite its adaptability, these plants are subject to diseases such as those caused by insects and fungus. Within this genus, Agave tequilana is primarily used for the elaboration of beverages and in soil conservation, but research also indicates this plant’s potential as a source of lignocellulosic bioenergy feedstocks. In this study, the authors describe the solution structure and biophysical characterization of two chitinases from A. tequilana as determined by small-angle X-ray scattering (SAXS) at the Stanford Synchrotron Radiation Lightsource in combination with theoretical structure prediction using Robetta. The low-resolution structures both exhibited two distinct domains connected by a linker region, and the theoretically predicted Rosetta structure showed consistency with the solution structures and could be docked into the SAXS envelopes. Interestingly, AtChi1 and AtChi2 both showed antifungal activities, suggesting that the Agave enzymes are involved in host defense mechanisms. Therefore, these enzymes could be considered as pathogenesis-related proteins with an important role in the plant’s defense mechanism against fungi.

In this study, Lopez et al. examined the uptake of As(III) versus As(V) by periphytic biofilms. They also fed these biofilms to larval mayflies. Both oxidation states of As were taken up by the biofilms, leading to ~6,000 times higher As concentrations relative to the initial solutions. However, the concentrations present in the larval mayflies were diluted relative to the biofilms (for both oxidation states), showing that As was not biomagnified in this food chain and suggesting that As was sequestered in the biofilm in a form that was not bioavailable. The authors used X-ray fluorescence imaging performed on beamline 2-3 at the Stanford Synchrotron Radiation Lightsource (SSRL) to examine the speciation of As in periphytic biofilms under their different experimental conditions. This showed that (1) As was associated with iron regardless of its initial oxidation state and (2) As(III) was oxidized to As(V) when taken up by the periphyton biofilm. The authors t concluded that the periphyton biofilm sequestered As by oxidizing As(III) to As(V), possibly via redox active iron or manganese oxides present within the biofilm. The As(V) then adsorbed strongly to iron oxides.

In summary, Lopez et al. have demonstrated the ability of benthic periphytic biofilms to accumulate As(V) and As(III) efficiently, without transferring As to higher trophic levels. This likely was because As was sequestered in a strongly adsorbed As(V) complex on iron oxide surfaces.

Sierra-Gomez, Y., et al. 2019. “A Biophysical and Structural Study of Two Chitinases from Agave tequilana and Their Potential Role as Defense Proteins,” The FEBS Journal 286(23), 4778−96. [DOI:DOI:10.1111/febs.14993.].