5UR2

Crystal structure of proline utilization A (PutA) from Bdellovibrio bacteriovorus inactivated by N-propargylglycine

Summary for 5UR2

• Entry DOI: 10.2210/pdb5ur2/pdb
• Descriptor: Bifunctional protein PutA, N-propargylglycine-modified flavin adenine dinucleotide (3 entities in total)
• Functional Keywords: flavoenzyme, rossmann fold, aldehyde dehydrogenase, flavin adenine dinucleotide, nicotinamide adenine dinucleotide, proline catabolism, substrate channeling, bifunctional enzyme, mechanism-based inactivation, oxidoreductase
• Biological source: Bdellovibrio bacteriovorus (strain ATCC 15356 / DSM 50701 / NCIB 9529 / HD100)
• Total number of polymer chains: 4
• Total formula weight: 442146.84

Authors

Tanner, J.J. (deposition date: 2017-02-09, release date: 2017-08-02, Last modification date: 2024-10-23)

Primary citation

Korasick, D.A.,Singh, H.,Pemberton, T.A.,Luo, M.,Dhatwalia, R.,Tanner, J.J.
Biophysical investigation of type A PutAs reveals a conserved core oligomeric structure.
FEBS J., 284:3029-3049, 2017

PubMed Abstract: Many enzymes form homooligomers, yet the functional significance of self-association is seldom obvious. Herein, we examine the connection between oligomerization and catalytic function for proline utilization A (PutA) enzymes. PutAs are bifunctional enzymes that catalyze both reactions of proline catabolism. Type A PutAs are the smallest members of the family, possessing a minimal domain architecture consisting of N-terminal proline dehydrogenase and C-terminal l-glutamate-γ-semialdehyde dehydrogenase modules. Type A PutAs form domain-swapped dimers, and in one case (Bradyrhizobium japonicum PutA), two of the dimers assemble into a ring-shaped tetramer. Whereas the dimer has a clear role in substrate channeling, the functional significance of the tetramer is unknown. To address this question, we performed structural studies of four-type A PutAs from two clades of the PutA tree. The crystal structure of Bdellovibrio bacteriovorus PutA covalently inactivated by N-propargylglycine revealed a fold and substrate-channeling tunnel similar to other PutAs. Small-angle X-ray scattering (SAXS) and analytical ultracentrifugation indicated that Bdellovibrio PutA is dimeric in solution, in contrast to the prediction from crystal packing of a stable tetrameric assembly. SAXS studies of two other type A PutAs from separate clades also suggested that the dimer predominates in solution. To assess whether the tetramer of B. japonicum PutA is necessary for catalytic function, a hot spot disruption mutant that cleanly produces dimeric protein was generated. The dimeric variant exhibited kinetic parameters similar to the wild-type enzyme. These results implicate the domain-swapped dimer as the core structural and functional unit of type A PutAs.

PubMed: 28710792
DOI: 10.1111/febs.14165

Experimental method

X-RAY DIFFRACTION (2.23 Å)