Since joining the lab in July, 1999 I have worked on several projects (some successful and some not). In the last couple of years I have been concentrating on two projects, 1) the structures of Cu,Zn superoxide dismutase from pathogenic organisms and 2) how mutations in human Cu,Zn superoxide dismutase result in Familial Amyotrophic Lateral Sclerosis (FALS). The latter project is being approached from structural, biochemical and biophysical perspectives.
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Cu,Zn Superoxide Dismutase (CuZnSOD)
Cu,Zn superoxide dismutase (CuZnSOD) is a dimeric enzyme consisting of two identical subunits of approximately 16 kDa each. Each monomer is composed of 8 antiparallel beta strands arranged in a Greek-key fold. Each monomer binds one copper and one zinc atom using 1 aspartic acid residue and 6 histidine residues. One histidine acts as a bridge between the metal sites ligating to both the copper and zinc atoms.
| Human SOD Dimer (PDB Code: 1SOS) | Human HSOD Active site |
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| Click image for rotating animation. (Animated GIF, ~ 1 MB) |
Click for larger Image. (800x600, JPEG) |
Reactive oxygen species (ROS) such as the superoxide and hydroxyl radicals are extremely harmful to biological macromolecules as they are able to oxidize lipids, modify proteins and cause DNA damage. SOD detoxifies superoxide radicals by converting them to hydrogen peroxide and water. Hydrogen peroxide is also toxic and it is eliminated through the action of catalase. SODs using other metal centers are also known, these include MnSOD, FeSOD and NiSOD. MnSOD is localized to the mitochondrial matrix in humans, while FeSOD and NiSOD are normally found in lower organisms.
Amyotrophic lateral sclerosis (ALS or Lou Gehrig's Disease) is a genetic, neurological disorder which affects approximately 1 in 200,000 people. The disorder which was first described by Jean-Martin Charcot in 1869, usually strikes in mid-life and results in the selective death of motor neurons leading to progressive paralysis and death within 1-5 years of onset of symptoms. In the vast majority of cases no genetic linkage has yet been identified. This form of the disorder is referred to as sporadic ALS (SALS). In 5-10% of cases a genetic linkage to mutations in CuZnSOD has been identified. This form of the disorder is referred to as famlilial ALS (FALS).
To date over 90 individual mutations have been identified in FALS patients. These mutations are dispersed throughout the protein and do not seem to be concentrated in any one specific region. The mechanism by which these mutation cause selective death of motor neurons is not understood. Current theories include 1) reduced or altered SOD activity leading to an increase in oxidative damage, 2) toxic accumulation of CuZnSOD containing aggregates resulting from protein misfolding or impaired protein degredation, 3) neurofilament disorganization leading to axonal strangulation and 4) excitotoxic death due to mishandling of glutamate. SOD aggregation is an attractive hyposthesis since protein aggregates are a common feature of other neurological disorder such as Alzheimer's, Huntington's and Prion diseases. However, why only motor neurons are selectively killed is still unknown.
Some current reviews on ALS:
Bacterial Cu,Zn SOD is also a homodimeric enzyme however its dimer interface is completely different from that used by the eukaryotic enzymes. The dimer interface in bacterial SODs uses a face of the beta-barrel which is directly across the beta-barrel from the face used in the eukaryotic enzyme. The bacterial dimer interface is also larger than its eukaryotic counterpart.
| Bacterial SOD Dimer (PDB Code: 2APS) |
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| Click image for rotating animation. (Animated GIF, ~ 750 kB) |
Cu,Zn SOD in bacteria is usually localized to the periplasmic space, and is utilized to protect the invading pathogen from the hosts immune response. When bacteria infect a host, the host's immune respose results in the generation of superoxide and other radicals designed to kill the invading pathogen. With high concentrations of SOD enzyme in their periplasmic space, bacterial pathogens are able to neutralize this flood of superoxide radicals thereby evading the host's immune system. Several studies have shown that SOD knockouts are approximately 1000 fold less pathogenic than wild-type strains.
©Michael DiDonato, PhD., The Scripps Research Institute
http://www.scripps.edu/~didonato/current.shtml
Last updated: Friday, 04-Jun-2004 14:44:17 PDT