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Characterization of tunnel mutants reveals a catalytic step in ammonia delivery by an aminoacyl‐ tRNA amidotransferase
Author(s) -
Zhao Liangjun,
Rathnayake Udumbara M.,
Dewage Sajeewa W.,
Wood Whitney N.,
Veltri Anthony J.,
Cisneros G. Andrés,
Hendrickson Tamara L.
Publication year - 2016
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1002/1873-3468.12347
Subject(s) - glutamine amidotransferase , deprotonation , ammonia , chemistry , protonation , residue (chemistry) , biochemistry , glutamine , mutant , transfer rna , amino acid , ion , organic chemistry , rna , gene
The Helicobacter pylori Asp‐ tRNA A sn /Glu‐ tRNA G ln amidotransferase (Gat CAB ) utilizes an uncommonly hydrophilic, ~ 40 Å ammonia tunnel for ammonia/ammonium transport between isolated active sites. Hydrophilicity of this tunnel requires a distinct ammonia transport mechanism, which hypothetically occurs through a series of deprotonation and protonation steps. To explore the initiation of this relay mechanism, the highly conserved tunnel residue D185 (in the GatA subunit) was enzymatically and computationally investigated by comparing D185A, D185N, and D185E mutant enzymes to wild‐type Gat CAB . Our results indicate that D185 acts as an acid/base residue, participating directly in catalysis. To our knowledge, this is the first example of acid/base chemistry in a glutamine‐dependent amidotransferase ammonia tunnel.