ISSN 0027-1314, Moscow University Chemistry Bulletin, 2018, Vol. 73, No. 2, pp. 80–84. © Allerton Press, Inc., 2018.
Original Russian Text © A.A. Pometun, S.Yu. Kleymenov, S.A. Zarubina, I.S. Kargov, P.D. Parshin, E.G. Sadykhov, S.S. Savin, V.I. Tishkov, 2018, published in Vestnik Moskovskogo
Universiteta, Seriya 2: Khimiya, 2018, No. 2, pp. 164–169.
Comparison of Thermal Stability of New Formate Dehydrogenases
by Differential Scanning Calorimetry
A. A. Pometun
, S. Yu. Kleymenov
, S. A. Zarubina
, I. S. Kargov
a, b, c
, P. D. Parshin
E. G. Sadykhov
, S. S. Savin
, and V. I. Tishkov
a, b, c,
Bach Institute of Biochemistry, Federal Research Center “Fundamentals of Biotechnology,”
Russian Academy of Sciences, Moscow, 119071 Russia
Innovations and High Technologies MSU Ltd., Moscow, 109559 Russia
Department of Chemistry, Moscow State University, Moscow, 119991 Russia
Koltzov Institute of Developmental Biology, Russian Academy of Sciences, Moscow, 119334 Russia
Received November 27, 2017
Abstract⎯Formate dehydrogenases (FDHs) from different sources are systematically studied in our labora-
tory. Over the past few years, new genes of four FDHs from pathogenic bacterium Staphylococcus aureus
(SauFDH), methylotrophic thermotolerant yeast Ogataea parapolymorpha (OpaFDH), yeast Saccharomyces
cerevisiae (SceFDH), and moss Physcomitrella patens (PpaFDH) have been cloned and expressed in E. coli
cells. By means of differential scanning calorimetry, a comparative study of thermal stability of new recombinant
formate dehydrogenases and a number of FDHs from other sources has been performed. It was shown that two new
enzymes, SauFDH and OpaFDH, are comparable to the FDH from Pseudomonas sp. 101 bacteria in their stability.
SceFDH is the least stable FDH among the described formate dehydrogenases.
Keywords: thermal stability, formate dehydrogenase, differential scanning calorimetry
Formate dehydrogenases (FDH, EC 18.104.22.168.) from
different sources have been studied in our laboratory
for many years [1–3]. FDH is present in bacteria,
yeast, and microscopic fungi, as well as in embryo-
phytes and nonvascular plants. The importance of
studying this enzyme is determined by the following
factors: FDH is actively used in practice as a biocata-
lyst for cofactor regeneration , and also plays an
important role in the vital functions of various organ-
isms [1, 3]. In order to evaluate the possibility of using
formate dehydrogenase as a biocatalyst, it is necessary
to obtain information about the kinetic parameters
and stability of this enzyme.
Genes encoding FDH in different organisms have
been successfully cloned in E. coli in many laboratories
all over the world. The creation of effective expression
vectors made it possible to obtain recombinant FDHs
in active and soluble forms. Our laboratory possesses
the world’s largest collection of cloned FDH genes.
This collection includes genes from bacteria Pseudo-
monas sp. 101 (PseFDH), Moraxella sp. C-1, and
Mycobacterium vaccae N10, methylotrophic yeast
Candida boidinii (CboFDH), and plants Arabidopsis
thaliana (AthFDH) and soya Gycine max (SoyFDH). In
recent years, we have cloned and expressed active forms of
new genes of FDH from pathogenic bacterium Staphylo-
coccus aureus (SauFDH), methylotrophic thermotolerant
yeasts Ogataea parapolymorpha (OpaFDH), baker’s yeasts
(SceFDH), and moss Physcomitrella patens (PpaFDH).
The study of the thermal stability of FDHs is an
important task, since many biocatalytic processes are
carried out for a long time at an elevated temperature.
As a rule, the thermal stability of formate dehydroge-
nases is studied by the following two methods: through
the inactivation kinetics and with differential scanning
calorimetry (DSC). In the present work, comparative
studies of the thermal stability of the four new and
some of the previously obtained recombinant FDHs
are performed by the DSC method.
Expression of Recombinant Formate Dehydrogenases
The expression of genes encoding the target for-
mate dehydrogenase was performed in the E. coli
BL21 (DE3) CodonPlus/pLysS cells. The producer
Abbreviations: DSC, differential scanning calorimetry;
SauFDH, PseFDH, OpaFDH, CboFDH, SceFDH, AthFDH,
SoyFDH, and PpaFDH, recombinant formate dehydroge-
nases extracted from bacteria Staphylococcus aureus and Pseu-
domonas sp. 101, methylotrophic yeasts Ogataea parapolymor-
pha and Candida boidinii, baker’s yeast, and plants A. thaliana,
Glycine max soya, and Physcomitrella patens, respectively.