A Study of Soluble Glucose Dehydrogenase Based upon Kinetic Studies of the Structural Analogs of PQQ.
James Ewing, Leslie Thompson & Carolnn Yong
Faculty Advisor: T. Otsuki

     Soluble glucose dehydrogenase is a homodimeric enzyme found in various bacteria such as Acinetobacter calcoaceticus, requiring Ca²⁺ and pyrroloquinolinequinone (PQQ) cofactors for full enzymatic activity. sGDH catalyses the oxidation of glucose and other aldose sugars. The mechanistic details have yet to be disclosed. Towards that goal, a specific series of 3 isomeric analogs (2-4) of PQQ are being synthesized from methoxyaniline derivatives in the laboratory. The synthesis includes the reaction of a diazonium salt with ethyl-2-methyacetoacetate to yield the corresponding hydrazones of ethyl pyruvate. Each hydrazone underwent acid catalyzed Fischer indole synthesis followed by reduction of the nitro group using H₂/Pd/C in to the corresponding aminoindole. Michael addition with dimethyl trans 2-ketoglutaconate was performed to form the third pyridine ring, which was further oxidized into triesters. Hydrolysis of the oxidation product yields the 3 isomeric analogs of PQQ.
     These isomers differ in the positioning of the nitrogen atom on the ring system and will serve to define structural significance in terms of binding with a metal cation and reconstitution process with the apoprotein. Detailed analysis of the stability of each reconstituted enzyme as well as steady-state kinetics studies will shed light on the structural characteristics mandatory for enzymatic activity and active site characterization.

Support provided by: National Science Foundation-REU, National Science Foundation-AIRE