BL 424 Chapter 11: Bioenergetics and Metabolism:

Mitochondria, Chloroplasts, Peroxisomes

In addition to involvement in protein sorting and transport, organelles are specialized
compartments for metabolism:
Mitochondria and chloroplasts are devoted to production of ATP.
Proteins for organelles are mostly synthesized on free ribosomes in the cytosol and
then directly transported; mitochondria and chloroplasts also have their own organelle
transcription and protein synthesis.

Student Learning outcomes:

Proteins are the active players in most cell processes.
1*. Explain concisely the similarities and differences in structure and function of these
organelles.
2*. Explain the process of transport of proteins to mitochondria and chloroplasts; to appreciate
the signals on proteins that identify their destinations..
3*. Describe the diverse protein complexes that read the protein’s signals and assist
the transport: TOM, TIM, TOC, TIC complexes, chaperones, peptidases.
4*. Explain the essential metabolic functions of mitochondria and chloroplasts, (of
oxidative phosphorylation and photosynthesis, respectively); note the importance of
membrane compartments for formation of proton gradients and for function of the
ATP synthase.
5. Describe the mitochondrial and chloroplast genomes, and that these
genes are transcribed and translated within the organelle.
6. Recall that prokaryotes complete oxidative phosphorylation and photosynthesis by
using their plasma membrane to generate the proton gradient.

Important Figures: 2*, 4*, 5*, 6, 7*, 10*, 12, 13, 14, 15*, 16*, 17*, 18, 22, 25*, 33
Important Tables: 1, 2

1*. Mitochondria structure and organization.

Mitochondria are critical for metabolic energy – for generation of ATP from the
breakdown of carbohydrates and fatty acids.

Mitochondria have a double membrane structure: (Fig. 11.1)

Matrix (interior) has enzymes of citric acid (TCA) cycle
to oxidize Acetyl CoA to CO2; also has DNA (Fig. 11.2)

Inner membrane has numerous proteins involved

in electron transport and oxidative phosphoyrlation (ATP).

Outer membrane is permeable to small molecules

Porins (channels) freely admit molecules < 1 kD.
Recall that pyruvate is imported from cytosol,
And converted to Acetyl CoA in matrix.
Mitochondria have DNA genomes: mostly circular, reflecting endosymbiotic origins.
Humans 16-kb; yeast 80-kb; plants > 200 kb.
Encode rRNAs, tRNAs and some proteins involved
in oxidative phosphorylation (about 13-32).

Ribosomes are in the matrix;

Translation in mitochondria has some special codon usage.

Mutations of mitochondrial genes cause human disease:

Mitochondria inherited from mother;
Usually mix of normal and mutant –
affect tissues with large ATP requirement (eye, brain)
LHON (Leber’s Hereditary Optic Neuropathy)

* Most mitochondrial proteins are encoded

by the nuclear genome: (Figs. 4-7).
About 1000 different proteins are synthesized
on free ribosomes (cytoplasm) and then imported.

Matrix proteins must traverse two membranes:

Positively-charged N-terminal presequences
(20-35 aa) target proteins for import;
Presequences are removed by peptidase
Matrix processing peptidase.

Tom complex (translocase of outer membrane)

includes receptors and channel proteins to
to transport across first membrane.
Tim complex (translocase of inner membrane)
carries proteins into the matrix (Fig. 11.4).
Chaperones assist the import, (Fig. 11.5, rachet)
ATP hydrolysis powers the movement.
Also electrical potential of membrane helps
import +++ presequence into matrix
(intermembrane space is more + charged H+ gradient)

Proteins destined for insertion in membranes: (Fig. 11.6)

Some transport proteins use Hsp90 and an
Internal sequence for import and insertion.
Lot of oxidative phosphorylation proteins in membranes
Other proteins can have presequences
and internal sequences (Fig. 11.7,8) to end up different places

Phospholipids are carried to mitochondria

from the ER by phospholipids transfer proteins.
Ex. Cardiolipin (4 fatty acids) in inner mitochondrial membrane
2. Mechanism of oxidative phosphorylation.
Most energy (ATP) from oxidative metabolism comes from the transfer of electrons from
NADH and FADH2 to a series of electron carriers in four complexes in the inner
mitochondrial membrane.

The transfer of electrons leads to proton transfer

across the membrane (establishing a proton gradient –
electrochemical gradient (Fig. 11.10*, 11.12*).

Chemiosmotic coupling hypothesis (Peter Mitchell):

(Fig. 11.12) A fifth protein complex, ATP synthase,
couples ATP synthesis to the return of protons
to the mitochondrial mateix (Fig. 11.13).
1 NADH -> 3ATP; 1 FADH2 -> 2 ATP.

The electrochemical, proton gradient also drives transport

of ATP, ADP and other metabolites into and out of the
Mitrochondrial matrix (Fig. 11.14).
(Outer membrane is permeable to small molecules).

3. Chloroplasts and other plastids

Chloroplasts are larger organelles (5-10 um; Fig. 11.15), and have 3 membranes:
the double membrane defines the organelle;
the stroma is equivalent to the mitochondrial matrix.
The internal thylakoid membrane is site of
electron transport and chemiosmotic ATP synthesis;
enzymes are located on outer (stromal) surface.

**Fig. 11.16 compares chemiosmotic generation of ATP

for mitochondrial and chloroplast compartments.

[Note: prokaryotes use their plasma membrane

for chemiosmotic ATP synthesis for photosynthesis
or oxidative phosphorylation.]

Chloroplast genomes are 120-160 kb,

and contain about 150 genes; several plant
chloroplast genomes have been sequenced.
Encode 4 rRNAs, 30 tRNAs and 21 ribosomal proteins,
plus ~30 proteins for photosynthesis (Table 2).
Some proteins are synthesized in chloroplast.
Import of proteins into chloroplast. About 95% of chloroplast proteins are encoded
by nuclear genes; synthesized on free ribosomes, and imported;
they must cross 2 or 3 membranes.

The target sequence is an N-terminal (30-100 aa)

Transit peptide which is bound by guidance complex
and directed to the Toc complex (Translocation
outer membrane chloroplast (Fig. 11.17).
Import requires Hsp70 chaperones, ATP, GTP hydrolysis.

The transit peptide is not positively charged, but the

intermembrane space (and stroma) do not differ in charge.

Transit across the inner membrane uses Tic complex

of receptors and channels; ATP is required;
the Hsp100 chaperone assists.
The transit peptide is cleaved in the stroma by
Stromal processing peptidase (SPP).

Proteins destined for the thylakoid lumen are

first imported into the stroma, and then a
second signal sequence is used for transport
across the thylakoid membrane (Fig. 11.18).

The N-terminal signals involve 3 different mechanisms:

Chaperones, + charged, and signal recognition particle.

*Other plant organelles (plastids) also have chloroplast genomes and two membranes,
but serve other functions. Plastids develop from small proplastids.

Chromoplasts lack chlorophyll

but have other pigments (carotenoids);
responsible for colors of flowers, vegetables.

Leucoplasts – nonpigmented, store energy sources:

Amyloplasts store starch (ex. potatoes)
Elaioplasts store lipids.

Chloroplasts require light for development:

In the dark, etioplasts with some internal
membranes are an intermediate state (Fig. 11.21).
4. Photosynthesis converts energy from the sun
to usable chemical energy form. Chlorophyll pigments
organized in photocenters absorb light; electrons
are excited and transferred to a series of carriers
in the thylakoid membranes: (Fig. 11.22; Fig. 11.25)
(photosystem II, photosystem I and cytochromes).

Transfer of electrons is coupled to formation

of proton gradient in the thylakoid lumen
(ATP synthesis by ATP synthase in the stroma,
with reduction of NADP+ to NADPH in stroma.

Electrons are obtained by hydrolysis of water

by photosystem II; photosystem I transfers
electrons to NADP reductase (Fig. 11.25).
[Cyclic electron flow uses electrons from
Photosystem I only, and generates extra ATP
but not NADPH (Fig. 11.27).]

5. Peroxisomes are small organelles with a single membrane that contain diverse
enzymes for fatty acid oxidation, glyoxylate cycle and photo-respiration (Fig. 11.28).
They often generate hydrogen peroxide; (Fig. 11.29)
they contain catalase to decompose peroxides.

Peroxisomes perform some biosynthesis of lipids

(cholesterol and dolichol in animal cells are
made here as well as in the ER).

Peroxisome assembly begins in ER with

formation of specific vesicles. Most peroxisome
proteins are synthesized on free ribosomes and
imported into nascent peroxisome (Fig. 11.33).
Two types of signals target proteins to peroxisome:
PTS1 is Ser-Lys-Leu at COOH end;
PTS2 is 9-aa sequence at N-terminal.
Signals are recognized by receptors and protein channels.

Human disease occurs from mutations in peroxisomal enzymes

or from defective import mechanisms.
(ex. Zellweger syndrome is mutated PTS1 receptor)
Lethal in 1st 20 years of life)

Review: Questions 1, 4, 5, 7, 8, 9, 11, 12.

Consider the experimental basis of protein transport, and determination of signals.
Compare the prokaryote model to that of organelles.