New chip finds the 'Factor X' in AIDS

If you wanted to show off the power of a new tool for identifying potentially therapeutic proteins, you couldn't do better than running a routine demonstration and chancing upon a long-sought Factor X that wards off AIDS.

It happened this year to US-based company Ciphergen Biosystems, which has developed a new protein-hunting technology called ProteinChip.

Cipherhgen's Dr Rebecca Caffrey described the find to last week's annual Australian Society of Microbiology (ASM) Conference 2002.

Caffrey's Ciphergen team, from Fremont, California, worked with researchers at Rockefeller University's Aaron Diamond AIDS Research Centre in New York, to unmask the mysterious factor called CAF, that has eluded AIDS researchers for 16 years.

CAF keeps the AIDS virus shackled and silent in a tiny minority of "long-term non-progressers" -- HIV-positive individuals who do have not progressed to full-blown AIDS years after being infected.

CAF was long known to be a soluble factor secreted by CD8 T-lymphocytes, the specialised hunter-killer immune-system cells that seek out and destroy virus-infected cells.

It turns out to be not one factor, but three tiny sibling proteins called alpha defensins, that have literally been falling off geneticists' protein gels.

Two-dimensional gel electroporesis has been the standard technique for sorting proteins for two decades. Protein extracts from biological samples are sorted by 'running' them through thin sheets of acrylamide gel, which separate them according to their specific combinations of charge and molecular weight.

Caffrey says defensins are tiny, highly charged proteins -- a combination that caused them literally to race through gels and fall off the edges.

Common wisdom held that anything with a molecular weight of less than 20 kilodaltons was probably peptide "junk" from the breakup of larger proteins; the region below 20 kilodaltons was regarded as a molecular equvalent of the Rub'al Khali -- nobody wanted to go there.

The three alpha-defensins that the US researchers have discovered all tip the molecular scale at less than 3.5 kilodaltons.

Caffrey is intrigued by their very high charge -- a characteristic of molecules that nature has designed to interact directly with DNA.

If her hunch is right, they may suppress replication of the AIDS virus in T-lymphocytes, macrophages and other susceptible immune-system cells by preventing replication of the virus' genetic blueprint -- that makes them potentially superb candidates for an anti-AIDS therapy.

Caffrey is also intrigued by the fact that there are three families of defensins in primates -- but humans make only alpha- and beta-defensins. They have the gene for the third group, the theta-defensins, but the gene has been inactivated.

Could a loss-of-function mutation in the human theta-defensin gene some time after the human-chimp divergence some 7 million years ago have left our species vulnerable to the HIV virus?

The HIV-virus is believed to have originated in chimps, which become HIV positive when exposed to the virus, but like long-term non-progressor human patients, do not develop full-blown AIDS.

New technology

Ciphergen's ProteinChip system is being sold in Australia and Asia by Melbourne-based biotech equipment dealer Advanced Labs.

ProteinChip employs a principle called surface-enhanced laser desorption and ionisation time-of-flight mass spectrometry, or SELDI-TOF.

Attached to a spectrometer, the device allows researchers to rapidly separate and catalogue the full spectrum of proteins in a biological sample. Richard Holder, Advanced Labs' managing director, says the exception sensitivity of the device allows detection of rare or unusual proteins that conventional techniques may have overlooked.

With SELDI-TOF, raw biological samples are placed in wells in small metal strips that have been coated with one of 10 different surfaces that selectively bind proteins with specific physico-chemical properties.

The unbound proteins are washed away, and the captured proteins are impregnated with an energy absorbing matrix compound. The chip is then placed in the ProteinChip device and zapped with a laser, causing the molecules to "fly" through the mass spectrometer -- the time of flight yields a measure of each protein's molecular weight.

Holder says ProteinChip does not replace gel systems, but augments them with its ability to detect proteins across the full spectrum of molecular weights between 1 and 200 kilodaltons.

Holder has already sold one of the $500,000 devices to Prof John Mattick's laboratory at the Institute of Molecular Bioscience at the University of Queensland.

During last week's ASM 2002 meeting, the company held talks with representatives of the Victorian government over the possibility of purchasing a ProteinChip device for Victorian biotech and biomedical researchers, and installing it as part of the research complex around Australia's first synchrotron near Monash University's Clayton campus.