Automated analytical platform facilitates identification of proteins

October 01, 2001

CHAMPAIGN, Ill. -- Now that the human genome has been sequenced, one of the hottest areas in life sciences is characterizing the human proteome. Researchers at the University of Illinois have developed techniques that facilitate the rapid identification and characterization of proteins.

"New analytical methodologies are needed to identify the form and function of the hundreds of thousands of proteins encoded by genes," said Neil Kelleher, a UI professor of chemistry. "Part of the problem is retrieving intact proteins from databases using high-resolution, tandem mass spectrometric data and correlating their predicted structures with those actually present in mature proteins."

Contemporary approaches to protein identification using mass spectrometry have involved the measurement of peptide masses, but the direct fragmentation of protein ions "can be far more efficient than exhaustive peptide mapping," Kelleher said. "This is a new strategy for proteome analysis."

Kelleher's instrumentation combines Fourier-Transform Mass Spectrometry with electrospray ionization and separation methods. At the heart of the system is a liquid-helium cooled superconducting magnet. A vacuum system and mass spectrometer extend into the magnet's center.

"This is a relatively new breed of magnet," Kelleher said. "Instead of using 12 tons of bulky steel, the magnet is actively shielded with a counter-propagating magnetic field. The fields cancel one another outside the magnet, but at the magnet's center the field strength is a hefty 9.4 tesla."

Fractionated proteins are squirted into the vacuum system and then transported into the magnet, where they begin to spin. "The proteins spin at different frequencies, depending on their mass and charge," Kelleher said. "We gradually excite their orbits to higher and higher radii, and they eventually fly past sensitive detector plates in the mass spectrometer."

Computers then analyze the data to identify and characterize the proteins. The entire system is becoming increasingly automated for ease and efficiency of operation.

For their initial studies, Kelleher and his students selected two representative life forms: Mycoplasma pneumoniae - a simple bacteria with a tiny genome - and Methanococcus jannaschii - an archaeon found in submarine hydrothermal vents.

First, the researchers showed that multiple proteins could be processed simultaneously. Then they tested a predictive model for database search specificity.

The model agreed well with actual searches from a database of about 3,500 protein forms predicted from the genomic sequence of M. jannaschii. The method also should work for the millions of possible protein forms predicted from the human genome.

"These conceptual and technical advances provide a powerful tool for protein characterization in the post-genomic era," Kelleher said. "By better characterizing proteins, we can improve our fundamental understanding of the blueprint of life." The researchers described their technique in the October issue of Nature Biotechnology.
-end-


University of Illinois at Urbana-Champaign

Related Human Genome Articles from Brightsurf:

240 mammals help us understand the human genome
A large international consortium led by scientists at Uppsala University and the Broad Institute of MIT and Harvard has sequenced the genome of 130 mammals and analysed the data together with 110 existing genomes to allow scientist to identify which are the important positions in the DNA.

The National Human Genome Research Institute publishes new vision for human genomics
The National Human Genome Research Institute this week published its 'Strategic vision for improving human health at The Forefront of Genomics' in the journal Nature.

Interpreting the human genome's instruction manual
Berkeley Lab bioscientists are part of a nationwide research project, called ENCODE, that has generated a detailed atlas of the molecular elements that regulate our genes.

3-D shape of human genome essential for robust inflammatory response
The three-dimensional structure of the human genome is essential for providing a rapid and robust inflammatory response but is surprisingly not vital for reprogramming one cell type into another.

The genome of chimpanzees and gorillas could help to better understand human tumors
A new study by researchers from the Institute of Evolutionary Biology (IBE), a joint center of UPF and the Spanish National Research Council (CSIC), shows that, surprisingly, the distribution of mutations in human tumors is more similar to that of chimpanzees and gorillas than that of humans.

It's in our genome: Uncovering clues to longevity from human genetics
Researchers from Osaka University found that high blood pressure and obesity are the strongest factors reducing lifespan based on genetic and clinical information of 700,000 patients in the UK, Finland and Japan.

New limits to functional portion of human genome reported
An evolutionary biologist at the University of Houston has published new calculations that indicate no more than 25 percent of the human genome is functional.

Synthesizing the human genome from scratch
For the past 15 years, synthetic biologists have been figuring out how to synthesize an organism's complete set of DNA, including all of its genes.

Science and legal experts debate future uses and impact of human genome editing in Gender & the Genome
Precise, economical genome editing tools such as CRISPR have made it possible to make targeted changes in genes, which could be applied to human embryos to correct mutations, prevent disease, or alter traits.

Evolution purged many Neanderthal genes from human genome
Neanderthal genetic material is found in only small amounts in the genomes of modern humans because, after interbreeding, natural selection removed large numbers of weakly deleterious Neanderthal gene variants, according to a study by Ivan Juric and colleagues at the University of California, Davis, published Nov.

Read More: Human Genome News and Human Genome Current Events
Brightsurf.com is a participant in the Amazon Services LLC Associates Program, an affiliate advertising program designed to provide a means for sites to earn advertising fees by advertising and linking to Amazon.com.