PDBsum entry 1w3n

Aldolase

PDB id: 1w3n

Contents

Protein chains

293 a.a.

Ligands

SSH ×4

GOL ×3

Waters ×812

References listed in PDB file

Key reference

Title

The structural basis for substrate promiscuity in 2-Keto-3-Deoxygluconate aldolase from the entner-Doudoroff pathway in sulfolobus solfataricus.

Authors

A.Theodossis, H.Walden, E.J.Westwick, H.Connaris, H.J.Lamble, D.W.Hough, M.J.Danson, G.L.Taylor.

Ref.

J Biol Chem, 2004, 279, 43886-43892. [DOI no: 10.1074/jbc.M407702200]

PubMed id

15265860

Abstract

The hyperthermophilic Archaea Sulfolobus solfataricus grows optimally above 80 degrees C and metabolizes glucose by a non-phosphorylative variant of the Entner-Doudoroff pathway. In this pathway glucose dehydrogenase and gluconate dehydratase catalyze the oxidation of glucose to gluconate and the subsequent dehydration of gluconate to D-2-keto-3-deoxygluconate (KDG). KDG aldolase (KDGA) then catalyzes the cleavage of KDG to D-glyceraldehyde and pyruvate. It has recently been shown that all the enzymes of this pathway exhibit a catalytic promiscuity that also enables them to be used for the metabolism of galactose. This phenomenon, known as metabolic pathway promiscuity, depends crucially on the ability of KDGA to cleave KDG and D-2-keto-3-deoxygalactonate (KDGal), in both cases producing pyruvate and D-glyceraldehyde. In turn, the aldolase exhibits a remarkable lack of stereoselectivity in the condensation reaction of pyruvate and D-glyceraldehyde, forming a mixture of KDG and KDGal. We now report the structure of KDGA, determined by multiwavelength anomalous diffraction phasing, and confirm that it is a member of the tetrameric N-acetylneuraminate lyase superfamily of Schiff base-forming aldolases. Furthermore, by soaking crystals of the aldolase at more than 80 degrees C below its temperature activity optimum, we have been able to trap Schiff base complexes of the natural substrates pyruvate, KDG, KDGal, and pyruvate plus D-glyceraldehyde, which have allowed rationalization of the structural basis of promiscuous substrate recognition and catalysis. It is proposed that the active site of the enzyme is rigid to keep its thermostability but incorporates extra functionality to be promiscuous.

Figure 3.
FIG. 3. S. solfataricus KGDA catalytic mechanism. Suggested mechanism of KDGA based on that proposed for NAL, indicating the ability of the enzyme to accept substrates that vary at both stereocenters C-4 and C-5. Complexes trapped in this study are shown in bold.

Figure 4.
FIG. 4. Substrate-enzyme interactions. Schematic summary of interactions made with the diastereomers KDG and KDGal.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2004, 279, 43886-43892) copyright 2004.