Picking Apart The Protein

Genomics continues to be all the rage in biotech circles—with special kudos to Illumina’s (San Diego, CA) recent ability to obtain an entire human genome sequence in 24-hours for a mere thousand dollars. Our overall understanding of human DNA combined with our ability to determine individual genomes leads to better disease insight, more powerful diagnostics, and a higher output of efficacious therapeutics.

Equally important, though not as well-understood, is the field of proteomics. While genomics testing lays out the odds of developing a particular disease, proteomics testing reveals the presence of disease and its stage of development. In this issue, we’ll pore over the proteome and prime our readers for this emerging biotech market.

Term Of The Week: Proteome & Proteomics

The proteome is the full complement of proteins produced—or, expressed—by the genes of a particular cell, tissue, or organism. Proteomics is the analysis of the structure, function, and interactions of those proteins. The big data generated by a proteomics study is organized into databases and applied to medical and biological questions.

Genome vs. Proteome

The genome is fixed for life—it’s the same at birth as it is at age 99. And that genome is the same in every cell of your body—this is why sequencing the genome of a white blood cell, for example, can yield genetic information about a patient’s liver. Of course, environmental exposures to certain chemicals or radiation may cause changes in the DNA of some cells, but for the most part, the genome is fixed.

The proteome varies from cell type to cell type. Even though a liver cell and a white blood cell have the same DNA sequence, each cell has unique characteristics and functions. Each cell type turns on and off distinct sets of genes and it is this feature that confers different cell characteristics.

The proteome is fluid over the course of a lifetime with constant change in protein production through the years—and in some cases throughout a day or hour. The amount of a specific protein often changes significantly between healthy and diseased states. Some illnesses are the result of an increase in the expression of proteins, such as the over-expression of HER2 in HER2-positive breast cancer. Other illnesses are the result of a decrease in the expression of proteins, such as the decreased expression of insulin in type 1 diabetes.

Importantly, these changes in the proteome can exist even when there are no differences in a diseased person’s genome in comparison to a healthy individual. Developing a better comprehension of the proteome in both diseased and healthy states is critical for truly understanding disease and developing new diagnostics.

Studying The Proteome: Mass Spectrometry

Determining the identity and relative concentrations of different proteins in biological samples can be technically challenging. One tried and true approach is mass spectrometry. This complex technology can be summarized as a series of steps:

1) Protein samples are treated so their complex three-dimensional structures are unfolded, making them easier to analyze.
2) Unfolded proteins are “digested” with enzymes that cut them up into smaller fragments called peptides.
3) The peptide fragments are “ionized,” or given an electrical charge. The process of ionization further fragments the peptides.
4) Positively charged peptide fragments move through a magnetic field towards a negatively charged panel.
5) Peptide fragments with different masses move at different rates, creating a graph or “mass spectrum” which is compared to the baseline spectrum in a database.
6) The comparison identifies the proteins present in the initial sample.

A number of companies have developed diagnostic tests based on variations of mass spectrometry: Biodesix’s (Boulder, CO) VeriStrat test measures the levels of different proteins in a blood sample of lung cancer patients to identify those with more aggressive cancers. The VeriStrat test also reveals which patients will respond best to Genentech’s (South San Francisco, CA) Tarceva. Applied Proteomics’ (San Diego, CA) SimpliPro analyzes eleven proteins correlated with colorectal cancer. Integrated Diagnostics’ (Seattle, WA) blood test determines the probability that a lung nodule is benign by measuring levels of multiple circulating proteins.

A Twist On Protein Microarrays

Protein microarrays are another way to determine the identity of proteins. Traditionally, antibodies are attached to a glass chip. Each antibody has an affinity for a specific protein. When samples are washed over the chip, proteins will attach to their respective antibodies. Once “captured,” the proteins are decoded.

SomaLogic (Boulder, CO) is bringing a new twist to the microarray game. Instead of using antibodies as the “capture” agent, they use DNA sequences called aptamers, which are selected in the lab for their affinity to specific proteins. These aptamer sequences are attached to microscopic beads and probed with the protein sample. The aptamer sequences that capture proteins can be easily quantified using techniques such as qPCR. SomaLogic markets an inflammation panel which determines the levels of sixteen inflammation-related proteins.

As the field of proteomics evolves, we can expect to see a more comprehensive understanding of disease translated into superior diagnostics and therapeutics.

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