ABSTRACT
Introduction...................................................................................................... 168 Organization of Mammalian Pyruvate Dehydrogenase Complex ................... 171 E2 and E3BP Domains and E1 and E3 Binding ......................................... 171 Inner Framework and Stoichiometry of E3BP to E3 Dimers ..................... 172
Specificity in the Use of Lipoyl Prosthetic Group and Lipoyl Domains in Component Reactions.............................................................................. 173 E1 Specificity ............................................................................................... 173 E2 and E3 Specificity .................................................................................. 174 Relative Use of Different Lipoyl Domains in Overall PDC and E3 Reactions ..................................................................................... 174
Regulatory Enzymes ........................................................................................ 175 Kinase Isoforms and Isoform Focus ............................................................ 175 PDK2 Structure ............................................................................................ 176 PDP Isoforms, PDP1 Subunits, and Focus.................................................. 177
Roles of Lipoyl Domains in the Regulation of Mammalian PDC.................. 177 Enhanced PDK and PDP1 Function ............................................................ 177 E2 Constructs ............................................................................................... 178 Regulatory Properties of PDK ..................................................................... 178
Regulated PDK2 Interaction with L2 .......................................................... 179 Effector Modification of PDK2 Activity and Structure............................... 179
Specific Nature of PDK3 Interaction with L2 ................................................. 180 Tight PDK3-L2 Complex ............................................................................ 180 Critical L2 Domain Structure for Binding and Activation of PDK3 .......... 181 L2 Prosthetic Group Structure for Binding and Activation of PDK3......... 184 Effects of Free Lipoyl Group Structures on PDK3 Activity and L2 Binding ........................................................................................ 185
Requirements for L2 Binding to PDP1 ........................................................... 186 Metal Requirements for PDP1 and PDP1c Binding to L2 Domain............ 186 L2 Domain Structure Needed for Binding PDP1c ...................................... 187
Conclusions...................................................................................................... 188 Acknowledgment ............................................................................................. 188 References ........................................................................................................ 188
The pyruvate dehydrogenase complex (PDC) catalyzes the irreversible conversion of pyruvate to acetyl-CoA and CO2 coupled to the reduction of NAD
þ to NADH. This overall reaction (Figure 7.1) results from sequential catalysis by the pyruvate dehydrogenase (E1), the dihydrolipoyl acetyltransferase (E2), and the dihydrolipoyl dehydrogenase (E3) components. The five-step overall reaction is integrated by the three central steps that use the lipoyl prosthetic group as a substrate. The substrate lipoyl group is attached to specific lysine residues on each among a large set of mobile lipoyl domains that serve as intermediate carriers moving between the active sites of E1, E2, and E3 (Patel and Roche, 1990; Reed and Hackert, 1990; Perham, 2000; Roche et al., 2001). Depending on the source, E2 subunits have one to three lipoyl domains. The capacity of the lipoyl domains for traversing efficiently between the E1, E2, and E3 active sites is advanced by the mobility of the extended linker regions that connect these and other globular domains in the E2 structures (Figures 7.1 and 7.2). The C-terminal inner (I) domains of the multidomain E2 subunits from a wide range of organisms assemble into either 24-subunit cubic structures or 60-subunit structures with the appearance of a pentagonal dodecahedron (Oliver and Reed, 1982; Reed and Hackert, 1990; Mattevi et al., 1992; Perham, 2000). Figure 7.2 shows a slab cut through the inner core of human E2. The lipoyl and component binding domains of E2 surround this inner core structure. Here, we will consider the roles of E2 in the organization and function of PDC with emphasis on the distinct features (and components) in mammalian PDC. We will consider specificity-determining lipoyl domain structure in PDC catalysis with emphasis on the E1 reaction.
