8PMR

NADase from Aspergillus fumigatus with mutated calcium binding motif (D219A/E220A)

Summary for 8PMR

• Entry DOI: 10.2210/pdb8pmr/pdb
• Related: 6YGE 6YGF 6YGG
• Descriptor: Conidial surface nicotinamide adenine dinucleotide glycohydrolase nadA, DI(HYDROXYETHYL)ETHER, 1,2-ETHANEDIOL, ... (12 entities in total)
• Functional Keywords: nadase, nad hydrolase, ca-binding, homodimer, glycoprotein, extracellular, tnt domain, hydrolase
• Biological source: Aspergillus fumigatus Af293
• Total number of polymer chains: 4
• Total formula weight: 116401.08

Authors

Kallio, J.P.,Ferrario, E.,Stromland, O.,Ziegler, M. (deposition date: 2023-06-29, release date: 2023-11-15, Last modification date: 2023-12-06)

Primary citation

Ferrario, E.,Kallio, J.P.,Stromland, O.,Ziegler, M.
Novel Calcium-Binding Motif Stabilizes and Increases the Activity of Aspergillus fumigatus Ecto-NADase.
Biochemistry, 62:3293-3302, 2023

PubMed Abstract: Nicotinamide adenine dinucleotide (NAD) is an essential molecule in all kingdoms of life, mediating energy metabolism and cellular signaling. Recently, a new class of highly active fungal surface NADases was discovered. The enzyme from the opportunistic human pathogen was thoroughly characterized. It harbors a catalytic domain that resembles that of the tuberculosis necrotizing toxin from , which efficiently cleaves NAD to nicotinamide and ADP-ribose, thereby depleting the dinucleotide pool. Of note, the NADase has an additional Ca-binding motif at the C-terminus of the protein. Despite the presence of NADases in several fungal divisions, the Ca-binding motif is uniquely found in the Eurotiales order, which contains species that have immense health and economic impacts on humans. To identify the potential roles of the metal ion-binding site in catalysis or protein stability, we generated and characterized NADase variants lacking the ability to bind calcium. X-ray crystallographic analyses revealed that the mutation causes a drastic and dynamic structural rearrangement of the homodimer, resulting in decreased thermal stability. Even though the calcium-binding site is at a long distance from the catalytic center, the structural reorganization upon the loss of calcium binding allosterically alters the active site, thereby negatively affecting NAD-glycohydrolase activity. Together, these findings reveal that this unique calcium-binding site affects the protein fold, stabilizing the dimeric structure, but also mediates long-range effects resulting in an increased catalytic rate.

PubMed: 37934975
DOI: 10.1021/acs.biochem.3c00360

Experimental method

X-RAY DIFFRACTION (1.94 Å)