discovering dry eye
The Chemistry of Proteins
BY JASON J. NICHOLS, OD, MPH, PHD, & KARI B. GREEN-CHURCH, PHD

The term "proteomics" is rather new and is intended to separate the science of understanding protein structure and function from genomics. The proteome is constantly changing and interacting with cells and other bio-molecules compared to a relatively stable genome.

Back to Basics

Polysaccharides, lipids and nucleic acids (DNA and RNA) are the primary biological macromole-cules composing living organisms. Nucleotides are the mono-
mers of nucleic acids, which are composed of phosphate group, pentose sugar and nitrogenous hetercyclous base (in DNA, purines include adenine and guanine while pyrimidines include thymine and cytosine). DNA is a polymer of these nucleotides encoding amino acids, and is often commonly referred to as the genetic code.

Amino acids, by definition, contain both an amino and carboxylic acid attached to a shared carbon, and there are 20 standard amino acids that are encoded (although many more amino acids have been identified, only two recently identified are genetically encoded). Endogenous processes synthesize some amino acids, while others are essential (they come from exogenous sources).

Finally, chains of amino acids form peptides, which in turn form proteins. Proteins are essential to the structure and function of all living cells, and are typically large molecules with masses up to 3,000,000 Daltons.

Assessing Proteins

A variety of methods exist to determine protein expression levels. The first step typically involves separation, followed by identification, quantification and sequencing. Traditional, gel-based methods in addition to SDS-PAGE and immunological methods (western blot and surface-enhanced laser desorption and Ionization [SELDI]) are still quite common for separation, but mass spectrometry has become a practical method for the quantitation and simultaneous identification of complex protein samples (such as those found in the tear film).

Proteins have many different functions. For example, many proteins are enzymes, molecules that catalyze biochemical reactions and are commonly denoted by terms ending in "-ase" preceded by the name of the molecule they modify (such as matrix metalloproteinases in the tear film). Many diseases are associated with over- or under-production of enzymes; researchers recently found that stimulation matrix metalloproteinase-9 is associated with a decrease in epithelial barrier function relating to dry eye and ocular surface desiccation.

Proteins also play structural and mechanical roles in the body. An ophthalmic example includes proteoglycans, which are glycosylated proteins (chondroitin sulfate) associated with collagen fibers in the corneal stroma. Collagen is known for its tremendous tensile strength, as is the cornea.

Another main function of proteins is their role in the immune response. For example, antibodies are proteins that help the body identify foreign material. Immunoglobulins (IgG, IgA, IgM, IgD, IgE) are antibodies produced by B cells of the humoral immune response. Portions of both IgG and IgM are associated with the tear film, as are other immune-related proteins such as cytokines (interleukins), tumor necrosis factor alpha and growth factors.

Pieces of the Puzzle

Proteins are important molecules to understand relative to unraveling the etiology of dry eye disease. We are still learning much about not only the individual proteins expressed in the tear film, but also their function and interaction with other molecules such as lipids. A basic understanding of these issues will contribute to developing new technologies and pharmaceutical strategies targeted toward treating this disease.

Dr. Nichols is assistant professor of optometry and vision science at The Ohio State University College of Optometry. Dr. Green-Church is the director of the Mass Spectrometry and Proteomics Facility at The Ohio State University.