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Title: Producing glucose 6-phosphate from cellulosic biomass: Structural insights into levoglucosan bioconversion

The most abundant carbohydrate product of cellulosic biomass pyrolysis is the anhydrosugar levoglucosan (1,6-anhydro-β-d-glucopyranose), which can be converted to glucose 6-phosphate by levoglucosan kinase (LGK). In addition to the canonical kinase phosphotransfer reaction, the conversion requires cleavage of the 1,6-anhydro ring to allow ATP-dependent phosphorylation of the sugar O6 atom. Using x-ray crystallography, we show that LGK binds two magnesium ions in the active site that are additionally coordinated with the nucleotide and water molecules to result in ideal octahedral coordination. To further verify the metal binding sites, we co-crystallized LGK in the presence of manganese instead of magnesium and solved the structure de novo using the anomalous signal from four manganese atoms in the dimeric structure. The first metal is required for catalysis, whereas our work suggests that the second is either required or significantly promotes the catalytic rate. Although the enzyme binds its sugar substrate in a similar orientation to the structurally related 1,6-anhydro-N-acetylmuramic acid kinase (AnmK), it forms markedly fewer bonding interactions with the substrate. In this orientation, the sugar is in an optimal position to couple phosphorylation with ring cleavage. We also observed a second alternate binding orientation for levoglucosan, and in these structures, ADP wasmore » found to bind with lower affinity. These combined observations provide an explanation for the high Km of LGK for levoglucosan. Furthermore, greater knowledge of the factors that contribute to the catalytic efficiency of LGK can be used to improve applications of this enzyme for levoglucosan-derived biofuel production.« less
Authors:
 [1] ;  [2] ;  [2] ;  [3] ;  [1] ;  [4] ;  [1]
  1. Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
  2. Michigan State Univ., East Lansing, MI (United States)
  3. Iowa State Univ., Ames, IA (United States)
  4. Univ. of Manitoba, Winnipeg, MB (Canada)
Publication Date:
OSTI Identifier:
1258283
Report Number(s):
LA-UR--15-27036
Journal ID: ISSN 0021-9258
Grant/Contract Number:
AC52-06NA25396
Type:
Accepted Manuscript
Journal Name:
Journal of Biological Chemistry
Additional Journal Information:
Journal Volume: 290; Journal Issue: 44; Journal ID: ISSN 0021-9258
Publisher:
American Society for Biochemistry and Molecular Biology
Research Org:
Los Alamos National Laboratory (LANL), Los Alamos, NM (United States)
Sponsoring Org:
USDOE
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 09 BIOMASS FUELS biofuel; carbohydrate processing; crystal structure; hexokinase; structural biology; anhydrosugar; levoglucosan