Incorporation of pyridoxal-5′-phosphate into the apoenzyme: A structural study of D-amino acid transaminase from Haliscomenobacter hydrossis

Incorporation of pyridoxal-5′-phosphate into the apoenzyme: A structural study of D-amino acid transaminase from Haliscomenobacter hydrossis Full article

Journal

Biochimica Et Biophysica Acta - Proteins and Proteomics
ISSN: 0926-6569 , E-ISSN: 1878-1454

Output data

Year: 2024, Volume: 1873, Article number : 141056, Pages count : DOI: 10.1016/j.bbapap.2024.141056

Authors

Bakunova Alina K ¹ , Matyuta Ilya O ¹ , Minyaev Mikhail E ² , Boyko Konstantin M ¹ , Popov Vladimir O ^1,3 , Bezsudnova Ekaterina Yu ¹

Affiliations

1	Bach Institute of Biochemistry, Research Centre of Biotechnology of the Russian Academy of Sciences, Moscow, Russia.
2	N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Moscow, Russia.
3	Department of Biology, Lomonosov Moscow State University, Moscow, Russia

Pyridoxal-5′-phosphate (PLP)-dependent transaminases are key enzymes of amino acid metabolism in cells and remarkable biocatalysts of stereoselective amination for process chemistry applications. As cofactor-dependent enzymes, transaminases are prone to cofactor leakage. Here we discuss the holoenzyme-apoenzyme interconversion and the kinetics of PLP incorporation into the apo form of a PLP-dependent transaminase from Haliscomenobacter hydrossis. PLP binding to the apoenzyme was slow in buffer, but was accelerated in the presence of substrates. Two crystal structures of the apoenzyme were obtained: the directly obtained apoenzyme (PDB ID: 7P8O) and the one obtained by soaking crystals of the holoenzyme in a phenylhydrazine solution (PDB ID: 8YRU). The mechanism of PLP association with the apoenzyme was proposed on the basis of structural analysis of these apo forms. Three rearrangement steps, including (I) anchoring of the PLP via the phosphate group, (II) displacement of two loops, and (III) Schiff-bonding between the PLP and the ε-amino group of the catalytic lysine residue, reconstituted the active holo form of the transaminase from H. hydrossis. The results obtained allowed us to determine in the active site a permanent part and elements that are assembled by PLP, these findings may be useful for transaminase engineering for biocatalysis.

Bakunova A.K. , Matyuta I.O. , Minyaev M.E. , Boyko K.M. , Popov V.O. , Bezsudnova E.Y.
Incorporation of pyridoxal-5′-phosphate into the apoenzyme: A structural study of D-amino acid transaminase from Haliscomenobacter hydrossis
Biochimica Et Biophysica Acta - Proteins and Proteomics. 2024. V.1873. 141056 . DOI: 10.1016/j.bbapap.2024.141056 WOS Scopus OpenAlex

≡ Web of science:	WOS:001339347700001
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