Transcript Final Poster- Mitochondrial carrier protein family

Fall, 2011. Final Semester Project
Biol 443, Biology and Biochemistry of Proteins
A Profile of the Mitto_carr Family
Kirsten West
INTRODUCTION
Proteins in the mitochondrial carrier
family are characterized as integral
membrane proteins that play
essential roles in the transport of
various metabolites across the inner
membranes of the mitochondrion.
Their most important role is in the
regeneration of ATP in eukaryotic cells
via oxidative phosphorylation. For
this process to take place, crossing of
the mitochondrial matrix is required
for both the uptake of the electron
rich substances ADP and inorganic
phosphate from the cytosol and for
the release of ATP into the cytosol.
STRUCTURE
Mitochondrial carriers are all made up of 6
transmembrane spanning a-helical
segments that form a dimer or two
identical subunits. A characteristic
tripartite structure is observed. This
structure is made up of 3 regions of
approximately 100 homologous amino acid
repeats that include 2 alpha helices and 1
large hydrophilic loop as well as the MCF
conserved motif. Both the N and C
terminus of the protein are located in the
intermembrane space.
ADP/ATP
transporter
This transport across the mitochondrial membrane is achieved
using an ADP/ATP transporter that functions to keep the
distribution of nucleotides equimolar on either side of the
mitochondrial membrane.
Interpro FAMILY DESCRIPTION
(IPR018108)
A variety of substrate carrier proteins that are
involved in energy transfer are found in the inner
mitochondrial membrane or integral to the
membrane of other eukaryotic organelles such as
the peroxisome [1, 2, 3, 4, 5, 6]. Such proteins
include: ADP, ATP carrier protein (ADP/ATP
translocase); 2-oxoglutarate/malate carrier
protein; phosphate carrier protein; tricarboxylate
transport protein (or citrate transport protein);
Graves disease carrier protein; yeast
mitochondrial proteins MRS3 and MRS4; yeast
mitochondrial FAD carrier protein; and many
others. Structurally, these proteins can consist of
up to three tandem repeats of a domain of
approximately 100 residues, each domain
containing two transmembrane regions.
PDB REPRESENTATIVE PROTEIN
PDB ID: 2C3E
PDB Entry Title: The Bovine Mitochondrial
ADP/ATP Carrier
Primary Citation Title: Structural basis for lipid
mediated interactions between mitochondrial
ADP/ATP carrier monomers
Authors: Nury, H., Dahout-Gonzalez, C.,
Trezeguet, V., Lauquin, G., Brandolin, G., PebayPeyroula, E.,
Journal: (2005) FEBS lett. 579: 6031
Structure of ADT1-BOVIN
Primary and Secondary Structures:
• 297 residues
• 70% helical
• 6 membrane-spanning alpha
helical segments
Tertiary Structure:
CATH Structure Classification
Class: Mainly Alpha
Architecture: Alpha/Alpha Barrel
ADT1-BOVIN has a three-fold
pseudo-symmetric structure,
consisting of a barrel of six
transmembrane α-helices with
three short α-helices on each of
the matrix loops and two loops in
the intermembrane space . The
odd numbered helices are
sharply kinked (20o to 35o) within
the membrane due to highly
conserved proline residues and
all other helices are tilted slightly
in relation to the membrane. The
positioning of the helices in the
membrane cause it to take on a
basket-like conformation that is
open to the intermembrane
space and closed to the matrix.
• Loop MI contains helices I and
II and h1-2, a very short helix
on the matrix loop
• Loop MII contains helices III
and IV and h3-4, a very short
helix on the matrix loop
• Loop MIII contains helices V
and VI, h5-6, and argenine
residues R234, R235, and R236
The Cavity
The basket like conformation of the
helices forms a large, mainly hydrophilic
cavity that is accessible from the
intermembrane space in the resting
conformation. The three argenine
residues R234, R235, and R236, of the
ADP/ATP carrier signature are all located
on the C-terminal end of helix 5. In
addition to the argenine residues, E264
is present on helix 5, forming a salt
bridge in complex with R236.
ACTIVE SITE
The active site of the ADT1-BOVIN is
located at the bottom of the cavity that
is formed by the proteins tripartite
arrangement of α- helices. All ADP/ATP
carriers consist of a triplication of a
sequence of 20 to 30 amino acid
residues located on the highly kinked
odd numbered α-helices
Argenine
(PxD/ExxK/RxK/R-(20 to 30 residues)- residue R234,
responsible for
D/EGxxxxaK/RG) and the unique
ATP binding
RRRMMM motif located at the C
terminus of helix 5. There are three
specific argenine residues involved in the
specificity of binding within the site,
R234, R235, and R236. Argenines R234
and R235 have side chains that are
Argenine residue
R236, responsible
accessible by the intermembrane space
for ADP binding.
for the binding of ATP, while the R236
Salt bridge
residue points towards the matrix and is formed with E264
shielded from the opening by a salt
bridge complex that it forms with E264.
When the transporter switches
conformations so that the cavity is then
open to the matrix, the R236 residue is
then responsible for the binding of ADP.
FUNCTION
The ADP/ATP carrier functions as a dimer and uses an antiport
mechanism that only binds adenine nucleotides that are not
complexed with magnesium. The carrier exports ATP from the
mitochondria in exchange for external ADP.
A cytoplasmic salt bridge network on the mitochondrial
membrane works in coordination with an argenine residue, on
kinked α- helix 5, to prevent conformational changes in the
absence of substrate.
The transporter is
said to function in a
‘pit’ to ‘channel’
conformation, where
opening is
dependent on the
binding of the
substrate to argenine
residues of the
conserved sequence
on the odd
numbered helices.
Once the substrate is bound, proline residues act as hinges,
straightening the kinked helices during the transport process to
open the pit into a channel.
ATP is bound to R234 and R235 first and forces the proline
residues to loosen the kink of the odd helices, opening the
channel. The ATP is transported out of the cell and the salt
bridge that was previously hiding the side chains of R236 and
E264 exposes these residues for ADP binding from the matrix.
The second nucleotide is transported into the cell and the
transporter is returned to its original conformation with the
cavity facing the intermembrane space.
The binding of both substrates takes place in close proximity
to the middle of the mitochondrial membrane.
Once a nucleotide is bound from either side of the carrier, it
favors binding of the second one from the opposite side,
preventing binding of two nucleotides from the same side on
each monomer. Consequently, the exchange of the substrates
will be equimolar and dependent on the presence of substrate
on either side of the membrane.
FAMILY STRUCTURE
Orthology
Group
Viruses
Bacteria
Archaea
#
Species
4
3
0
Eukaryota
527
Fungi
130
70
Viridiplantae
Metazoa
229
The Mitochondrial Carrier
protein is not seen in
Archaea. The protein is scarce
among bacteria and viruses
but seen readily in plants,
fungi, and especially animals
Paralogy
Paralogous proteins are homologous proteins, visible in one species,
that have diverged from one another through gene duplication
events or gene mutations by various means.
The following is a list of paralogues to mitochondrial carriers found
in the human genome:
Database: NCBI Protein Reference Sequences
11,008,086 sequences; 3,852,209,289 total letters
Query=
>2C3E:A|PDBID|CHAIN|SEQUENCESDQALSFLKDFLAGGVAAAISKTAVAPIERVKLLLQVQHASKQISAEKQYKGIIDCVVRIPKEQG
FLSFWRGNLANVIRYFPTQALNFAFKDKYKQIFLGGVDRHKQFWRYFAGNLASGGAAGATSLCFVYPLDFARTRLAADVGKGAAQREFT
GLGNCITKIFKSDGLRGLYQGFNVSVQGIIIYRAAYFGVYDTAKGMLPDPKNVHIIVSWMIAQTVTAVAGLVSYPFDTVRRRMMMQSGR
KGADIMYTGTVDCWRKIAKDEGPKAFFKGAWSNVLRGMGGAFVLVLYDEIKKFV
Length=297
Sequences producing significant alignments:
Value
Score(Bits)
E
ref|NP_001142.2| ADP/ATP translocase 1 [Homo sapiens]
585 0.0
ref|NP_001627.2| ADP/ATP translocase 3 [Homo sapiens]
558 0.0
ref|NP_001143.2| ADP/ATP translocase 2 [Homo sapiens]
557 0.0
ref|NP_112581.1| ADP/ATP translocase 4 [Homo sapiens]
444 2e-161
ref|NP_037518.3| calcium-binding mitochondrial carrier protei... 130 2e-37
ref|NP_998816.1| calcium-binding mitochondrial carrier protei... 130 2e-37
ref|NP_689920.1| graves disease carrier protein [Homo sapiens] 118 7e-34
ref|NP_077008.2| calcium-binding mitochondrial carrier protei... 117 2e-32
ref|NP_003696.2| calcium-binding mitochondrial carrier protei... 115 3e-31
ref|NP_848621.2| solute carrier family 25 member 42 [Homo sap... 111 3e-31
ref|NP_055066.1| calcium-binding mitochondrial carrier protei... 112 2e-30
ref|NP_001153682.1| calcium-binding mitochondrial carrier pro... 112 2e-30
ref|NP_001006644.1| calcium-binding mitochondrial carrier pro... 109 2e-30
ref|NP_775908.2| solute carrier family 25 member 41 [Homo sap... 108 8e-30
ref|NP_660348.2| solute carrier family 25 member 43 [Homo sap... 107 9e-30
ref|NP_001006642.1| calcium-binding mitochondrial carrier pro... 109 1e-29
ref|NP_443133.2| calcium-binding mitochondrial carrier protei... 108 2e-29
ref|NP_001006643.1| calcium-binding mitochondrial carrier pro... 108 2e-29
ref|NP_113669.1| mitochondrial glutamate carrier 2 [Homo sapi... 104 8e-29
ref|NP_001010875.1| kidney mitochondrial carrier protein 1 [H... 104 8e-29
BIBLIOGRAPHY & DATABASES
• BLASTP 2.2.26+ Reference: Stephen F. Altschul, Thomas L. Madden, Alejandro A. Schaffer, Jinghui Zhang, Zheng Zhang, Webb
Miller, and David J. Lipman (1997), "Gapped BLAST and PSI-BLAST: a new generation of protein database search programs",
Nucleic Acids Res. 25:3389-3402.
• The Pfam protein families database: R.D. Finn, J. Mistry, J. Tate, P. Coggill, A. Heger, J.E. Pollington, O.L. Gavin, P.
Gunesekaran, G. Ceric, K. Forslund, L. Holm, E.L. Sonnhammer, S.R. Eddy, A. BatemanNucleic Acids
Research (2010) Database Issue 38:D211-222
• http://www.ebi.ac.uk/interpro/DisplayIproEntry?ac=IPR018108
• Edmund R.S Kunji, The role and structure of mitochondrial carriers, FEBS Letters, Volume 564, Issue 3, 30 April 2004, Pages
239-244, ISSN 0014-5793, 10.1016/S0014-5793(04)00242-X.
(http://www.sciencedirect.com/science/article/pii/S001457930400242X) Keywords: Mitochondrial carrier; Electron
crystallography; X-ray crystallography; Structure; Membrane protein
• F. Palmieri, Mitochondrial carrier proteins, FEBS Letters, Volume 346, Issue 1, 6 June 1994, Pages 48-54, ISSN 0014-5793,
10.1016/0014-5793(94)00329-7.(http://www.sciencedirect.com/science/article/pii/0014579394003297) Keywords: Carrier
protein; Transmembrane topology; Bacterial expression; Transport; Liposome; Mitochondrion
• John E. Walker, The mitochondrial transporter family, Current Opinion in Structural Biology, Volume 2, Issue 4, August 1992,
Pages 519-526, ISSN 0959-440X, 10.1016/0959-440X(92)90081-H.
(http://www.sciencedirect.com/science/article/pii/0959440X9290081H)
• Lucy R. Forrest, Reinhard Krämer, Christine Ziegler, The structural basis of secondary active transport mechanisms,
Biochimica et Biophysica Acta (BBA) - Bioenergetics, Volume 1807, Issue 2, February 2011, Pages 167-188, ISSN 0005-2728,
10.1016/j.bbabio.2010.10.014. (http://www.sciencedirect.com/science/article/pii/S000527281000722X) Keywords:
Secondary active transport; Protein structure; Carrier; Coupling; Modeling; Protein conformation
• Title: Relations between structure and function of the mitochondrial ADP/ATP carrier
Author(s): Nury H. ; Dahout-Gonzalez C. ; Trezeguet V. ; et al.
Source: ANNUAL REVIEW OF BIOCHEMISTRY Book Series: Annual Review of Biochemistry
• Biophysics: Yi Wang and Emad Tajkhorshid; Electrostatic funneling of substrate in mitochondrial inner membrane
carriersPNAS 2008 ; published ahead of print July 8, 2008,doi:10.1073/pnas.0801786105