9B1V

Crystal structure of PqqT with PQQ and Gd3+ bound

9B1V の概要

• エントリーDOI: 10.2210/pdb9b1v/pdb
• 分子名称: Putative ABC transporter periplasmic solute-binding protein, 1,2-ETHANEDIOL, PYRROLOQUINOLINE QUINONE, ... (5 entities in total)
• 機能のキーワード: periplasmic binding protein, transport protein
• 由来する生物種: Methylorubrum extorquens
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 33779.44

構造登録者

Boggs, G.D.,Bruchs, A.T.,Thompson, P.J.,Olshansky, L.,Bridwell-Rabb, J. (登録日: 2024-03-13, 公開日: 2024-08-21)

主引用文献

Thompson, P.J.,Boggs, D.G.,Wilson, C.A.,Bruchs, A.T.,Velidandla, U.,Bridwell-Rabb, J.,Olshansky, L.
Structure-driven development of a biomimetic rare earth artificial metalloprotein.
Proc.Natl.Acad.Sci.USA, 121:e2405836121-e2405836121, 2024

PubMed Abstract: The 2011 discovery of the first rare earth-dependent enzyme in methylotrophic AM1 prompted intensive research toward understanding the unique chemistry at play in these systems. This enzyme, an alcohol dehydrogenase (ADH), features a La ion closely associated with redox-active coenzyme pyrroloquinoline quinone (PQQ) and is structurally homologous to the Ca-dependent ADH from the same organism. AM1 also produces a periplasmic PQQ-binding protein, PqqT, which we have now structurally characterized to 1.46-Å resolution by X-ray diffraction. This crystal structure reveals a Lys residue hydrogen-bonded to PQQ at the site analogously occupied by a Lewis acidic cation in ADH. Accordingly, we prepared KA- and KD-PqqT variants to assess the relevance of this site toward metal binding. Isothermal titration calorimetry experiments and titrations monitored by UV-Vis absorption and emission spectroscopies support that KD-PqqT binds tightly ( = 0.6 ± 0.2 μM) to La in the presence of bound PQQ and produces spectral signatures consistent with those of ADH enzymes. These spectral signatures are not observed for WT- or KA-variants or upon addition of Ca to PQQ ⸦ KD-PqqT. Addition of benzyl alcohol to La-bound PQQ ⸦ KD-PqqT (but not Ca-bound PQQ ⸦ KD-PqqT, or La-bound PQQ ⸦ WT-PqqT) produces spectroscopic changes associated with PQQ reduction, and chemical trapping experiments reveal the production of benzaldehyde, supporting ADH activity. By creating a metal binding site that mimics native ADH enzymes, we present a rare earth-dependent artificial metalloenzyme primed for future mechanistic, biocatalytic, and biosensing applications.

PubMed: 39116128
DOI: 10.1073/pnas.2405836121

実験手法

X-RAY DIFFRACTION (1.55 Å)