energetic barrier [67]. After the polymerization the microtubule stores the curvature
elastic energy, which seems to be utilized in the depolymeization process accompanying the restoration of the curved protofilaments [61] and also in the chromosomal
movement [67–69]. One would expect that the seam plays some special role in the
initiation of the depolymerization, but this point remains to be studied [57].
7.6.2.2 The Role of GTP in Microtubule Polymerization
Each tubulin binds one GTP molecule. As shown in Fig. 7.16, the GTP molecule
bound to alpha tubulin is “covered” by the beta tubulin, and remains bound to the
same site as GTP. Thus, the GTP binding site of alpha tubulin is called N
(non-exchangeable)-site. On the other hand, the GTP bound to the beta tubulin is
hydrolyzed to GDP by catalytic action of the side chain of Glu254 of alpha tubulin in
the adjacent heterodimer in the same protofilament. The GDP thus formed
exchanges with the solution GTP after the heterodimer dissociates from the microtubule. Thus, the GTP binding site on beta tubulin is called E (exchangeable)-site.
In vivo, the energy liberated by the hydrolysis of GTP is ~30 kJ/mol [59]. But the
polymerization does not require hydrolysis of GTP, because heterodimer binding
non-hydrolyzable GTP analog, GMP-CPP, can polymerize [67]; if the beta tubulin
binds GDP at E-site, the heterodimer is not polymerizable. The GTP hydrolysis in
microtubule dynamics has been related to the change in the intrinsic curvature of a
tubulin heterodimer. As a result of hydrolysis at the E-site, majority of heterodimers
in a microtubule bind GDP at the inter-heterodimer interface. Thus, the outward
curvature of the protofilament becomes apparent during the depolymerization (the
protofilaments exhibiting outward curvature before they fall apart [61]). This curvature should stem from structural features of the heterodimer; it may reflect the
intrinsic curvature of heterodimer binding GDP, or may be specific to the
heterodimer without interaction with adjacent heterodimers in the microtubule
lattice. A study has shown that the latter is the case [69]. The mode of the interaction
between heterodimers seems to be essential for the curvature of the protofilament. A
polymerized microtubule stores an elastic energy.
Upon polymerization of a heterodimer, GTP bound to the E-site is hydrolyzed, as
described above. Like actin polymerization, when the heterodimer concentration is
low, GTP hydrolysis occurs at the same rate as the polymerization of heterodimer,
but when the heterodimer concentration is high, GTP hydrolysis lags behind the
polymerization. As a result, the end of microtubule is occupied by a GTP-cap, the
accumulation of heterodimers binding GTP on beta subunit. It has been suggested
that the length of the cap can be as small as one layer of heterodimer [66]. The GTP
cap plays a central role in the dynamics of polymerization/depolymerization (see
Sect. 7.6.4). A study using antibodies against the tubulin that binds GTP has
demonstrated that majority of the antibodies binds to the growing end of
microtubules [70].
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elastic energy, which seems to be utilized in the depolymeization process accompanying the restoration of the curved protofilaments [61] and also in the chromosomal
movement [67–69]. One would expect that the seam plays some special role in the
initiation of the depolymerization, but this point remains to be studied [57].
7.6.2.2 The Role of GTP in Microtubule Polymerization
Each tubulin binds one GTP molecule. As shown in Fig. 7.16, the GTP molecule
bound to alpha tubulin is “covered” by the beta tubulin, and remains bound to the
same site as GTP. Thus, the GTP binding site of alpha tubulin is called N
(non-exchangeable)-site. On the other hand, the GTP bound to the beta tubulin is
hydrolyzed to GDP by catalytic action of the side chain of Glu254 of alpha tubulin in
the adjacent heterodimer in the same protofilament. The GDP thus formed
exchanges with the solution GTP after the heterodimer dissociates from the microtubule. Thus, the GTP binding site on beta tubulin is called E (exchangeable)-site.
In vivo, the energy liberated by the hydrolysis of GTP is ~30 kJ/mol [59]. But the
polymerization does not require hydrolysis of GTP, because heterodimer binding
non-hydrolyzable GTP analog, GMP-CPP, can polymerize [67]; if the beta tubulin
binds GDP at E-site, the heterodimer is not polymerizable. The GTP hydrolysis in
microtubule dynamics has been related to the change in the intrinsic curvature of a
tubulin heterodimer. As a result of hydrolysis at the E-site, majority of heterodimers
in a microtubule bind GDP at the inter-heterodimer interface. Thus, the outward
curvature of the protofilament becomes apparent during the depolymerization (the
protofilaments exhibiting outward curvature before they fall apart [61]). This curvature should stem from structural features of the heterodimer; it may reflect the
intrinsic curvature of heterodimer binding GDP, or may be specific to the
heterodimer without interaction with adjacent heterodimers in the microtubule
lattice. A study has shown that the latter is the case [69]. The mode of the interaction
between heterodimers seems to be essential for the curvature of the protofilament. A
polymerized microtubule stores an elastic energy.
Upon polymerization of a heterodimer, GTP bound to the E-site is hydrolyzed, as
described above. Like actin polymerization, when the heterodimer concentration is
low, GTP hydrolysis occurs at the same rate as the polymerization of heterodimer,
but when the heterodimer concentration is high, GTP hydrolysis lags behind the
polymerization. As a result, the end of microtubule is occupied by a GTP-cap, the
accumulation of heterodimers binding GTP on beta subunit. It has been suggested
that the length of the cap can be as small as one layer of heterodimer [66]. The GTP
cap plays a central role in the dynamics of polymerization/depolymerization (see
Sect. 7.6.4). A study using antibodies against the tubulin that binds GTP has
demonstrated that majority of the antibodies binds to the growing end of
microtubules [70].
118
7 Moving Life