Cell-biological assessment of human glucokinase mutants causing maturity-onset diabetes of the young type 2 (MODY-2) or glucokinase-linked hyperinsulinaemia (GK-HI)
Cv. Burke et al., Cell-biological assessment of human glucokinase mutants causing maturity-onset diabetes of the young type 2 (MODY-2) or glucokinase-linked hyperinsulinaemia (GK-HI), BIOCHEM J, 342, 1999, pp. 345-352
Mutations in the glucokinase (GK) gene cause type-2 maturity-onset diabetes
of the young type 2 (MODY-2) and GK-linked hyperinsulinaemia (GK-HI). Reco
mbinant adenoviruses expressing the human wild-type islet GK or one of four
mutant forms of GK, (the MODY-2 mutants E70K, E300K and V203A and the GK-H
I mutant V455M) were transduced into glucose-responsive insulin-secreting b
eta-HC9 cells and tested functionally in order to initiate the first analys
is in vivo of recombinant wild-type and mutant human islet GK. Kinetic anal
ysis of wild-type human GK showed that the glucose S-0.5 and Hill coefficie
nt were similar to previously published data in vitro (S-0.5 is the glucose
level at the half-maximal rate). E70K had half the glucose affinity of wil
d-type, but similar enzyme activity. V203A demonstrated decreased catalytic
activity and an 8-fold increase in glucose S-0.5 when compared with wild-t
ype human islet GK. E300K had a glucose S-0.5 similar to wild-type but a 10
-fold reduction in enzyme activity. E300K mRNA levels were comparable with
wild-type GK mRNA levels, but Western-blot analyses demonstrated markedly r
educed levels of immunologically detectable protein, consistent with an ins
tability mutation. V455M was just as active as wild-type GK, but with a mar
kedly reduced S-0.5. The effects of the different GK mutants on glucose-sti
mulated insulin release support the kinetic and expression data. These expe
riments show the utility of a combined genetic, biochemical and cell-biolog
ical approach to the quantification of functional and structural changes of
human GK that result from MODY-2 and GK-HI mutations.