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Structural and functional studies on Salmonella typhimurium pyridoxal kinase: the first structural evidence for the formation of Schiff base with the substrate
Author(s) -
Deka Geeta,
Kalyani Josyula N.,
Jahangir Fathima Benazir,
Sabharwal Pallavi,
Savithri Handanahal S.,
Murthy Mathur R. N.
Publication year - 2019
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.14933
Subject(s) - schiff base , pyridoxal , salmonella , chemistry , biochemistry , substrate specificity , substrate (aquarium) , enzyme , microbiology and biotechnology , stereochemistry , biology , bacteria , genetics , ecology
A large number of enzymes depend on the ubiquitous cofactor pyridoxal 5′ phosphate ( PLP ) for their activity. Pyridoxal kinase ( PLK ) is the key enzyme involved in the synthesis of PLP from the three forms of vitamin B 6 via the salvage pathway. In the present work, we determined the unliganded structure of St PLK in a monoclinic form and its ternary complex with bound pyridoxal ( PL ), ADP and Mg 2+ in two different tetragonal crystal forms (Form I and Form II). We found that, in the ternary complex structure of St PLK , the active site Lys233 forms a Schiff base linkage with the substrate ( PL ). Although formation of a Schiff base with the active site Lys229 was demonstrated in the Escherichia coli enzyme based on biochemical studies, the ternary complex of St PLK represents the first crystal structure where the Schiff bond formation has been observed. We also identified an additional site for PLP binding away from the active site in one of the ternary complexes (crystal Form I), suggesting a probable route for the product release. This is the first ternary complex structure where the modeled γ‐phosphate of ATP is close enough to PL for the phosphorylation of the substrate. St PLK prefers PL over pyridoxamine as its substrate and follows a sequential mechanism of catalysis. Surface plasmon resonance studies suggest that St PLK interacts with apo‐ PLP –dependent enzymes with μ m affinity supporting the earlier proposed direct transfer mechanism of PLP from PLK to PLP ‐dependent enzymes.