posted on 2020-06-09, 00:13authored byKyriacos Petratos, Renate Gessmann, Vangelis Daskalakis, Maria Papadovasilaki, Yannis Papanikolau, Iason Tsigos, Vassilis Bouriotis
The
structure of a recombinant (His-tagged at C-terminus) alcohol
dehydrogenase (MoADH) from the cold-adapted bacterium Moraxella sp. TAE123 has been refined with X-ray diffraction
data extending to 1.9 Å resolution. The enzyme assumes a homo-tetrameric
structure. Each subunit comprises two distinct structural domains:
the catalytic domain (residues 1–150 and 288–340/345)
and the nucleotide-binding domain (residues 151–287). There
are two Zn2+ ions in each protein subunit. Two additional
zinc ions have been found in the crystal structure between symmetry-related
subunits. The structure has been compared with those of homologous
enzymes from Geobacillus stearothermophilus (GsADH), Escherichia coli (EcADH), and Thermus sp. ATN1
(ThADH) that thrive in environments of diverse temperatures.
Unexpectedly, MoADH has been found active from 10
to at least 53 °C and unfolds at 89 °C according to circular
dichroism spectropolarimetry data. MoADH with substrate
ethanol exhibits a small value of activation enthalpy ΔH‡ of 30 kJ mol–1. Molecular
dynamics simulations for single subunits of the closely homologous
enzymes MoADH and GsADH performed
at 280, 310, and 340 K showed enhanced wide-ranging mobility of MoADH at high temperatures and generally lower but more
distinct and localized mobility for GsADH. Principal
component analysis of the fluctuations of both ADHs resulted in a
prominent open–close transition of the structural domains mainly
at 280 K for MoADH and 340 K for GsADH. In conclusion, MoADH is a very thermostable,
cold-adapted enzyme and the small value of activation enthalpy allows
the enzyme to function adequately at low temperatures.