Protein FAQ

Exons are nucleotide sequences in DNA and RNA that are conserved in the creation of mature RNA. The process by which DNA is used as a template to create mRNA is called transcription. mRNA then works in conjunction with ribosomes and transfer RNA (tRNA), both present in the cytoplasm, to create proteins in a process known as translation. Exons usually include both the 5’- and 3’- untranslated regions of mRNA, which contain start and stop codons, in addition to any protein coding sequences.

Introns are nucleotide sequences in DNA and RNA that do not directly code for proteins, and are removed during the precursor messenger RNA (pre-mRNA) stage of maturation of mRNA by RNA splicing. Introns can range in size from 10’s of base pairs to 1000’s of base pairs, and can be found in a wide variety of genes that generate RNA in most living organisms, including viruses. Four distinct types of introns have been identified:

Absolutely it can, and in at least two ways. Firstly, as the oncometabolites that we have been discussing, as an early warning indicator for cancer. Secondly, by interfering with cancer cell D-amino acid production. We think that cancer cells actually produce D-amino acids to protect themselves against certain cancer drugs. So they are fighting back and they can do this in a matter of minutes to hours. Nobody realized this before but, knowing this, we can develop drugs to interfere with D-amino acid production by cancer cells. This will make other cancer drugs more effective.

I think it may be the best forum or venue for a large number of both academic and industrial scientists to interact and communicate. In addition, the students are there and it is the best opportunity for them to hear your work and for you to hear about what they are doing, as well.

Yes, they can. In cancer, some of the amino acids appear to be upregulated, meaning they are in a higher concentration than you would normally find. Three in particular, D-aspartic acid, D-serine and D-alanine, are agonists for NMDA (N-methyl-D-aspartate) receptors. These are ion channels that are found upregulated in nerve tissues, the brain and in cancer cells as well. Neurologically, a D-amino acid has been found to stop severe seizures, and others are neurotransmitters, and some are agonists to neurotransmitters. We are not entirely sure as to the entire mechanism(s) of action, because we are just now finding out that these things exist.

Enantiomeric separations by HPLC is by far the most important technique. Mass spectrometry is more or less useless for qualitative analysis because enantiomerics have exactly the same mass, but it is useful as a sensitive detector for HPLC.

The nice thing about being an academic is that I can investigate anything I find to be interesting, regardless of the field. One would have to have a fairly limited imagination to focus on only one thing your entire life. Discoveries in one area usually impact other areas, so a new analytical method can be used in many fields. Once you invent a new method or a technique, even the most limited researchers can and do use it in many important fields. It is the invention of the new effective technique that mattered.

The main protein or peptide in Alzheimer's disease patients is something called beta-amyloid. It has a 42 amino acid chain, and accumulates in the plaque between nerve cells in Alzheimer's disease patients. Beta-amyloid has been found to epimerize. Certain amino acids epimerize in the beta-amyloid at specific locations, and it turns out those amino acids are mainly aspartic acid and serine. There was no good way to analyze for such aberrations in beta-amyloid until now, and we have found a number of techniques that not only detect it, but can point out exactly where, within the beta-amyloid chain, the aberration is occurring and if there are more aberrations in Alzheimer's patients than in normal people. Image Credit:Shutterstock/Tavarius

This work is going to expand greatly. We are just at the very beginning and are still trying to convince people that it is important and something that should be studied. I think that battle is over, and now the great onslaught of research is about to begin. The roles of D-amino acids, whether they are free D-amino acids or in peptides and proteins, more and more are going to be discovered. This research is going to expand greatly. I think you will find a field, D-amino acid proteomics, which we are proposing, is going to grow tremendously.

I am going to be talking about new columns based on ionic liquids. An ionic liquid is a liquid in which all the components are ions, like salt (sodium chloride), only, instead of being a solid, it is a liquid. These things have very high polarity and can be developed with extremely high thermal stabilities. They are highly useful as GC stationary phases, which I will be talking about. They are really useful as headspace solvents for the analysis of environmental and biological samples. We use it to analyze water and all sorts of materials. That is the thrust of what I will be talking about, new ionic liquids that are highly polar, highly stable and can be used for unusual analyses that ordinary stationary phases and ordinary techniques have difficulty with.

All living systems are made up of L-amino acids – we will call them left-handed amino acids - and D-amino acids are a mirror image of these. We will call those the right-handed amino acids. As with your right and left hand, one is a non-superimposable mirror image to the other. D-amino acids were originally thought to be unnatural and useless. We now know this is not the case. We are finding that they play important roles in a variety of biological processes. Image Credit: Shutterstock/ Christoph Burgstedt

Techniques are always evolving and the areas that they are going to be used in are going to be virtually unlimited. Petrochemistry, for example, is just one area, although that is the area of this particular talk. They are just as useful in environmental analysis, food and beverage analysis, fragrance, forensic science. It cuts across all these areas, so the research involving ionic liquids, or using them, can go in any direction.

It is one of the largest gathering of analytical and other scientists, and yet, all the presentations are in a relatively localized area, so one can easily access a greater variety of topics and scientists in a limited amount of time. It is also one of the biggest exhibitions in the world, and having this exhibition can provide useful technical information, often unknown or unexplained in academic talks. I bring a significant number of students to Pittcon. They must give a talk or poster in order to attend. I think this is really important as it is good for them. They love the whole experience, from the networking, giving their presentations, being exposed to new topics, new parts of the country. A lot of them have not traveled so it is a wonderful opportunity. That is why we always bring as many students as we can and have them participate; they love it.

We found that certain amino acids are elevated in cancer cells, relative to non-tumorigenic cells. This raises the question of why? This also means that they can be used as oncometabolites, which is simply something that is elevated in a cancer cell or a cancer patient, relative to what it is in a healthy person. As a diagnostic test, you can go in and if you find this oncometabolite, it is an indication that there is a cancer that needs to be examined or looked for a little more closely. So, it can be used as an early warning system, which is important.

We recently proposed that there is a possible test as a malignancy indicator using the ratio of certain D and L-amino acids, which seemed to work in the limited cell growth media that we were working in. However, it needs to be tested further and in more detail before any clinical applications are forthcoming. Image Credit:Shutterstock/CI Photos

As in the past, I hope the new technologies and new knowledge will be useful in the future. It is also important because, in doing my research, the next generation of scientists are educated and can continue such work long after I am gone. That is how science advances, often dragging humanity up and along with it.

Pittcon offers ease of networking, with a very large number of scientists in many different fields, all together in one location. It is easier to hear a wider variety of talks than at any other meeting I have been to, so the number of people you can access and talk to is much easier at Pittcon than virtually any meeting worldwide.

Gas chromatography and liquid chromatography are the dominant techniques for analysis of most things; gas chromatography if they are volatile, liquid chromatography if they are not. They are used for both the identification and quantification of materials, including very complex mixtures. When you get into the real world environment, the biological samples, these are very complex mixtures. It is interesting that the main source of employment in all areas of chemistry involves separations. If you do separations, then you will have a job. While employment opportunities were reduced during the pandemic, we did not experience that. Our students were getting job offers right through COVID, even with remote interviewing.

S. aureus is a Gram-positive and cocci-shaped bacterium, meaning that it appears purple with ‘Gram stain’, and these bacterial are seen as clusters of small, spherical cells. S. aureus is a part of the human microbiome – mostly being found on the surface of the skin and all mucous membranes (e.g. eyes, mouth and ears). They are opportunistic bacteria, meaning that if a break in the skin occurs, or if the host becomes immunodeficient in some way, S. aureus can eventually cause an infection. When S. aureus does infect a human host, it may result in a variety of different diseases, but very commonly it causes recurrent soft tissue and skin infections. These infections have become more prevalent in recent years, mainly as nosocomial forms, as methicillin-resistant strains of S. aureus (MRSA) have evolved in hospitals across the western world. In humans, protein A (once it is released into the extra-cellular environment) can bind with the ‘Fc’ fragment of immunoglobulin G (IgG) and has been observed to be antiphagocytic in vitro - which is consistent with its IgG binding abilities, as binding to IgG can interfere with bacterial attachment by the antibody. This means that it can prevent itself from being destroyed by certain elements of the human immune system (as was also similarly observed in the guinea pig experiment from 2013). S. aureus infections, such as endocarditis, are usually treated with a strong course of antibiotics. Protein A can therefore function slightly differently in different species, but always has the same goal: it interferes with the host’s B-cells within their immune system to prevent S. aureus from being phagocytosed and destroyed. This is an ideal adaptation, and when coupled with the known propensity of S. aureus to be more easily transmitted than other coagulase-positive bacteria, it indicates faster and more widespread bacterial transmission.

The main function of Protein A within the cell wall of S. aureus bacteria involves the host’s immune system. It is a virulence determinant (a molecule which determines how easily a pathogen can cause disease in a host) and plays a role in suppressing host B-cell responses - which therefore assists in preventing the host immune response from damaging the bacteria. Protein A also binds to the ‘Fc region’ of IgG, and also to the ‘Fab regions’ of B-cells – triggering processes which ultimately block opsonophagocytosis, and kills the B-cells that it comes in contact with. An experiment performed in 2015 showed that guinea pigs which were purposely infected with the S. aureus bacterial strain called SpAKKAA showed increased anti-S. aureus immune response. Immunisation with the SpAKKAA strain can be used to elicit the production of neutralizing antibodies – enabling the guinea pigs to develop sufficient protective immunity.