title: Binding and channeling of alternative substrates in the enzyme DmpFG: a molecular dynamics study.
abstract: DmpFG is a bifunctional enzyme comprised of an aldolase subunit, DmpG, and a dehydrogenase subunit, DmpF. The aldehyde intermediate produced by the aldolase is channeled directly through a buried molecular channel in the protein structure from the aldolase to the dehydrogenase active site. In this study, we have investigated the binding of a series of progressively larger substrates to the aldolase, DmpG, using molecular dynamics. All substrates investigated are easily accommodated within the active site, binding with free energy values comparable to the physiological substrate 4-hydroxy-2-ketovalerate. Subsequently, umbrella sampling was utilized to obtain free energy surfaces for the aldehyde intermediates (which would be generated from the aldolase reaction on each of these substrates) to move through the channel to the dehydrogenase DmpF. Small substrates were channeled with limited barriers in an energetically feasible process. We show that the barriers preventing bulky intermediates such as benzaldehyde from moving through the wild-type protein can be removed by selective mutation of channel-lining residues, demonstrating the potential for tailoring this enzyme to allow its use for the synthesis of specific chemical products. Furthermore, positions of transient escape routes in this flexible channel were determined.
authors.affiliation: Research School of Biology, Australian National University, Canberra, Australian Capital Territory, Australia.
authors.forename: Natalie E
authors.initials: NE
authors.lastname: Smith
authors.affiliation: School of Chemistry and Biochemistry, University of Western Australia, Perth, Western Australia.
authors.forename: Alice
authors.initials: A
authors.lastname: Vrielink
authors.affiliation: School of Chemistry and Biochemistry, University of Western Australia, Perth, Western Australia.
authors.forename: Paul V
authors.initials: PV
authors.lastname: Attwood
authors.affiliation: Research School of Biology, Australian National University, Canberra, Australian Capital Territory, Australia. Electronic address: ben.corry@anu.edu.au.
authors.forename: Ben
authors.initials: B
authors.lastname: Corry
mesh_terms: D000447:Aldehydes / Q000737:chemistry / Q000378:metabolism; D020134:Catalytic Domain; D005634:Fructose-Bisphosphate Aldolase / Q000737:chemistry / Q000378:metabolism*; D007651:Keto Acids / Q000737:chemistry / Q000378:metabolism; D056004:Molecular Dynamics Simulation; D009154:Mutation / Q000235:genetics; D010088:Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013816:Thermodynamics
chemical_list: D000447:Aldehydes; D007651:Keto Acids; D021122:Protein Subunits; D010088:Oxidoreductases; D005634:Fructose-Bisphosphate Aldolase
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