The document discusses biomolecular motors, particularly focusing on proteins that convert ATP hydrolysis into mechanical force, essential for various cellular functions like movement and muscle contraction. It details flagellar motors in bacteria and ATP synthase's structure and function, highlighting their roles in energy production and cellular transport mechanisms. Key components, mechanisms of action, and related gene functions are also outlined, emphasizing the complexity and significance of these molecular machines in biological systems.

1. Biomolecular Motors
Presented By:
Arushe Tickoo
DTU, Delhi

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The typical biomolecular motor is a protein that uses free energy from ATP (Adenosine Tri-
phosphate) or other nucleotide triphosphate (NTP) hydrolysis as its fuel to produce mechanical
force.
ATP is the universal currency of energy in biological systems and, is generated from glucose by
a sequence of reactions called glycolysis.
The hydrolysis of ATP to ADP and inorganic phosphate is energetically favorable and the pool of
ATP in a cell is far higher than that of its hydrolysis products, hence serving as a remarkable
store of chemical energy that is used to perform the various cellular functions.
Motor proteins are involved in several critical cellular processes and have functions ranging
from ATP synthesis and muscle contraction.
Biomolecular motor

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Flagellated bacteria, such as Escherichia coli, swim by rotating thin helical filaments, each driven at its
base by a reversible rotary motor, powered by an ion flux. A motor is about 45 nm in diameter and is
assembled from about 20 different kinds of parts. It develops maximum torque at stall but can spin several
hundred Hz.
Its direction of rotation is controlled by a sensory system that enables cells to accumulate in regions
deemed more favorable.
No more than 50 nm in Diameter It spins clockwise (CW) orcounterclockwise (CCW) at speeds on the order of
100 Hz, driving long thin helical filaments that enable cells to swim Receptors near the surface of the cell count
molecules of interest (sugars, amino acids, dipeptides) and control the direction of flagellar rotation. If a leg of
the search is deemed favorable, it is extended,
Flagellar motor

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Fig; A schematic diagram
of the flagellar motor
CheY-P
It is the chemotaxis
signaling molecule that
binds to FliM, and FlgM
is the anti-sigma factor
pumped out of the cell by
the transport apparatus;

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A cell swims steadily in a direction roughly parallel to its long axis for about a second—it is said to
“run”—and then moves erratically in place for a small fraction of a second—it is said to “tumble”—
and then swims steadily again in a new direction. When a cell runs at top speed, all of its flagellar
filaments spin CCW, the filaments form a bundle that pushes the cell steadily forward. When a cell
tumbles, one or more filaments spin CW; these filaments leave the bundle, and the cell changes
course
Motors switch from CCW to CW and back again approximately at random. The likelihood of
spinning CW is enhanced by a chemotactic signaling protein, CheY.
When phosphorylated, CheY binds to the cytoplasmic face of the flagellar motor. The
phosphorylation of CheY is catalyzed by a kinase, the activity of which is controlled by
chemoreceptors.
The different components of the motor are named after the genes that encode them. genes for which
mutant cells lacked flagellar filaments were called fla (for flagellum), but after more than 26 had been
found.
fla genes are now called flg, flh, fli, or flj, depending upon their location on the genetic map. Genes for
which mutant cells produce paralyzed flagella are called mot (for motility). Four of the all gene
products that are involved in gene regulation (FlgM, FlhC, FlhD, FliA);

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The M-ring (for membrane) has affinity for inner-
membrane fractions,
The S-ring is seen just above the inner membrane,
The P-ring (for peptidoglycan) is at the right place to be
embedded in the peptidoglycan,
and the L-ring (for lipopolysaccharide) has affinity for
outer-membrane fractions.
It was found that both the M- and S-rings (now called the
MS-ring) comprise different domains of the same protein,
FliF. Therefore, they function as a unit.

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FliG, FliM, and FliN are also referred to as the “switch complex,” since many mutations of fliG, fliM,
and fliN lead to defects in switching (in control of the direction of rotation)
Operons encoding the proteins of the chemotaxis system of E.
colia
Class 1 Class 2 Class 3
flhDC flgAMN fliC
flgBCDEFGHIJKL motABcheAW
flhBAE tar tap cheRBYZ
fliAZY aer
fliDST trg
Class 1 contains the master operon, flhDC, the expression of which is required for transcription of class 2 and class 3
operons.
Class 2 contains eight operons that encode components required for construction of the hook-basal body complex,
and class 3 contains six more that encode components required for filament assembly and motor function

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ATP Synthase
ATP synthase is one of the wonders of the molecular world. ATP synthase is an enzyme, a molecular motor, an
ion pump, and another molecular motor all wrapped together in one amazing nanoscale machine. It plays an
indispensable role in our cells, building most of the ATP that powers our cellular processes.
Human mitochondrial (mt) ATP synthase, or complex V consists of two functional domains: F1, situated in the
mitochondrial matrix, and Fo, located in the inner mitochondrial membrane. Complex V uses the energy created
by the proton electrochemical gradient to phosphorylate ADP to ATP.
ATP synthase consists of two well defined protein entities: the F1 sector, a soluble portion situated in the
mitochondrial matrix, and the Fo sector, bound to the inner mitochondrial membrane. F1 is composed of three
copies of each of subunits α and β, and one each of subunits γ, δ and ε. F1 subunits γ, δ and ε constitute the
central stalk of complex V. Fo consists of a subunit c-ring (probably comprising eight copies, as shown in
bovine mitochondria and one copy each of subunits a, b, d, F6 and the oligomycin sensitivity-conferring
protein (OSCP). Subunits b, d, F6 and OSCP form the peripheral stalk which lies to one side of the complex.

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Human mitochondrial ATP synthase, or
complex V, consists of two functional domains,
F1 and Fo. F1 comprises 5 different subunits
(three α, three β, and one γ, δ and ε) and is
situated in the mitochondrial matrix. Fo contains
subunits c, a, b, d, F6, OSCP and the accessory
subunits e, f, g and A6L. F1 subunits γ, δ and ε
constitute the central stalk of complex V.
Subunits b, d, F6 and OSCP form the peripheral
stalk. Protons pass from the intermembrane
space to the matrix through Fo, which transfers
the energy created by the proton
electrochemical gradient to F1, where ADP is
phosphorylated to ATP.
Structure of ATP Synthase
Jonckheere I, Smeitink J A M, and Rodenburg R J; Mitochondrial ATP synthase: architecture, function and
pathology (2011), J Inherit Metab Dis

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Fo Region F1 Region
The F1 portion is soluble and consists of a
hexamer, denoted a3b3. This hexamer is arranged
in an annulus about a central shaft consisting of the
coiled-coil γ subunit.
The Fo portion consists of three transmembrane
subunits: a, b2 and c10-14. The remainder of Fo
consists of the transmembrane subunits a, and
b2; the latter is attached by the d subunit to the
a3b3 hexamer so that it anchors the a subunit
to F1. Thus there are two ‘stalks’ connecting Fo
to F1: ge and b2d.

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The binding change mechanism. Notation for site occupancies: T = ATP bound, DP = ADP • Pi bound, D =
ADP bound. b subunits are numbered clockwise. The length of the arrows indicates the relative binding
affinities (a) The system starts with either (β 1, β 2, β 3) = (E, T D•P, D) (b) Clockwise rotation of g
increases the binding affinity of ADP in b1, traps ATP in b2, and promotes Pi binding on b3. (c) Further
rotation of g traps ADP and allows Pi binding in b1, releases the tightly bound ATP and allows ADP
binding in b2, and traps Pi in b3.

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In steps 1 and 2 a site binds ADP and phosphate (not necessarily in that order). While trapped in the catalytic
site in step 3, reactants (ADP and Pi ) and product (ATP) are in chemical equilibrium. Step 4 requires the
input of mechanical torque from Fo on g to trap the reactants in the ATP state and to pry open the site
releasing the tightly bound ATP. The way in which this works is found in the shape of the a3b3 hexamer and
the g shaft
At the top of the a3b3 hexamer is a hydrophobic ‘sleeve’ in which the g shaft rotates. Further down, however,
the annulus is offset from the center, so that as g rotates clockwise, it sequentially pushes outwards on each
catalytic site. In addition, the e subunit is located eccentrically and attached to the g and c subunits so that, as
g rotates, it comes into contact sequentially with each b subunit in a conserved region called the DELSEED
sequence (named for the single letter abbreviation of its constituent amino acids). Together, this asymmetric
rotation exerts stress on the catalytic site loosening its grip on ATP so that thermal fluctuations can free it into
solution.

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A single molecule of F1-ATPase acts as a rotary motor, the smallest known, by direct observation
of its motion. A central rotor of radius approximately 1 nm, formed by its gamma-subunit, turns
in a stator barrel of radius approximately 5nm formed by three alpha- and three beta-subunits. F1-
ATPase, together with the membrane-embedded proton-conducting unit F0, forms the H+-ATP
synthase that reversibly couples transmembrane proton flow to ATP synthesis/hydrolysis in
respiring and photosynthetic cells. In the presence of ATP, the filament rotated for more than 100
revolutions in an anticlockwise direction when viewed from the 'membrane' side. The rotary
torque produced reached more than 40 pN nm(-1) under high load.
Direct observation of the rotation of F1-ATPase.
Noji H, Yasuda R, Yoshida M, Kinosita K Jr.

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In the realm of ATP-driven linear biomotors, the most prominent examples are dynein, kinesin and
myosin, which is involved in muscle contraction. Muscle contraction is a sophisticated process involving
actin and myosin and is a result of actin filaments sliding on myosin heads. Actin filaments are formed of
375 amino acid long subunits that are associated with ATP. Myosin transport along actin is not limited to
muscle contraction, and is involved in several cellular transport processes.
It is thought that the two heads of the myosin move along in a hand-over-hand mechanism based on the
~74nm displacements observed by fluorescent labeling methods.
Interaction of an actin filament
with myosin-coated surface

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1. Itoh, H., et al., (2004) Mechanically driven ATP synthesis by F1-ATPase. Nature,. 427(6973): p. 465-8.
2. Noji, H., et al., (1997) Direct observation of the rotation of F1-ATPase. Nature,. 386(6622): p. 299-302.
3. Oster G, Wang H; ATP Synthase: Two rotary molecular motors working together; University of
California, Berkeley
4. Antoniel M etal., (2014 May) The Oligomycin-Sensitivity Conferring Protein of Mitochondrial ATP
Synthase ; Int J Mol Sci.; 15(5): 7513–7536.
5. Oster G and Wang H; (April 1999) ATP synthase: two motors, two fuels, Elsevier Science
6. Manuela Antoniel, (2014 May) The Oligomycin-Sensitivity Conferring Protein of Mitochondrial ATP
Synthase: Emerging New Roles in Mitochondrial Pathophysiology, International Journal of Molecular
Sciences
References