Stochastic boundary molecular dynamics simulation of L-ribose in the active site of Actinoplanes missouriensis xylose isomerase and its Val135Asn mutant with improved reaction rate.
We used molecular dynamics simulations to study how a non-natural substrate, L-ribose, interacts with the active site of Actinoplanes missouriensis xylose isomerase. The simulations showed that L-ribose does not stay liganded in the active site in the same way as D-xylose, in which the oxygens O2 and O4 are liganded to the metal M1. The oxygen O4 of L-ribose moved away from the metal M1 to an upside down position. Furthermore, the distances of the carbons C1 and C2 of L-ribose to the catalytic metal M2 were higher than in the case of D-xylose. These findings explain the extremely low reaction rate of xylose isomerase with L-ribose. The mutation V135N close to the C5-OH of the substrate increased the reaction efficiency 2- to 4-fold with L-ribose. V135N did not affect the reaction with D-xylose and L-arabinose, whereas the reaction with D-glucose was impaired, probably due to a hydrogen bond between Asn-135 and the substrate. When L-ribose was the substrate, Asn-135 formed a hydrogen bond to Glu-181. As a consequence, O4 of L-ribose stayed liganded to the metal M1 in the V135N mutant in molecular dynamics simulations. This explains the decreased K(m) of the V135N mutant with L-ribose.
Santa
Harri
H
Laboratory of Bioprocess Engineering, Helsinki University of Technology, P.O. Box 6100, 02015-TKK, Finland.
Kammonen
Juha
J
Lehtonen
Olli
O
Karimäki
Johanna
J
Pastinen
Ossi
O
Leisola
Matti
M
Turunen
Ossi
O
D019747:Aldose-Ketose Isomerases / Q000737:chemistry* / Q000235:genetics*; D019943:Amino Acid Substitution; D001665:Binding Sites / Q000235:genetics; D008848:Micromonosporaceae / Q000201:enzymology*; D015394:Molecular Structure; D009154:Mutation; D012266:Ribose / Q000737:chemistry*
D012266:Ribose; D019747:Aldose-Ketose Isomerases; C023298:xylose isomerase