A New Tool for Predicting Protein Aggregation

Many human disorders, such as Alzheimer’s and Parkinson’s, are characterized by misfolded proteins that aggregate (combine). It is important to understand the aggregation of proteins when creating soluble protein-based drugs. Therefore, methods that can predict the properties of protein aggregation are essential. Within the past decade, various tools have been developed to anticipate protein aggregation. Ventura et. al developed AGGRESCAN, which is the first tool that has relied on experimental patterns of aggregation on a cellular level. Mutants of the amyloid β-peptide (Aβ) were used to create the algorithm. Aggregation in the cytoplasm of E. coli was studied. Each mutation affects the peptide aggregation differently. The various aggregation patterns can be used to determine the likelihood of aggregation for the natural amino acids. AGGRESCAN is capable of using this information to determine the tendency of aggregation for each protein based on its relative position in the sequence. The algorithm has been commonly used due to its accuracy in predicting in vivo aggregation.

            The majority of the methods used to predict protein aggregation analyze linear sequences. Hence, they assume that the protein of interest is partially unstructured. For folded globular proteins, these regions may be blocked and therefore do not have a significant impact on aggregation. The authors introduce AGGRESCAN3D (A3D), which has evolved from the AGGRESCAN method. A3D allows the aggregation properties of globular proteins to be predicted because it is able to detect spatial relationships. A3D includes a “Dynamic Mode” that can be used, which considers the different structures that the input structure can have in the predicted aggregation patterns. Therefore, when wild type structures and destabilizing pathogenic mutations change in structure, A3D can account for the variations and model the aggregation patterns that stem from those variations in structure.

            A3D is documented online on a server and can be used to predict the likelihood of aggregation for globular proteins. The algorithm can be used to anticipate the effect of mutations in conformational disorders. It can also be used to design soluble protein-based drugs that can be used as treatment for disorders characterized by protein aggregation. Therefore, it provides the possibility of more effective treatment for patients with disorders like Alzheimer’s and Parkinson’s.

Reference:

R. Zambrano, M. Jamroz, A. Szczasiuk, J. Pujols, S. Kmiecik, S. Ventura. “AGGRESCAN3D (A3D): server for prediction of aggregation properties of protein structures”. Nucleic Acids Research, 2015; DOI: https://doi.org/10.1093/nar/gkv359

Cold Sores: A Complete Annoyance

Have you ever had a cold sore/fever blister? If you have, then you know how painful and annoying they can be. Cold sores consist of small blisters on and/or around the lips. The area is usually red, sore and even itchy. They usually heal within a few days, and there are many medications available that speed up the healing process.

Cold sores are caused by the herpes simplex virus 1 (HSV1). Once infected, the virus remains dormant in the body until it is triggered by a stressor, such as stress, fatigue, excessive sunlight, or a weakened immune system. Cold sores are very common, for approximately 80 % of the U.S. population has been exposed to this virus. I’ve always wondered why some of us consistently have cold sores while others never get them. Those lucky people have an immune system that is able to suppress the virus.

I and other cold-sore sufferers would love it if we never had to deal with another cold sore.  Researchers at Duke University may just make this a reality one day.  They have determined how the virus remains hidden. They now understand how the virus switches from the latent phase to the active phase on a molecular level. Using this information, they may be able to reawaken the virus and kill it.

For example, while HSV1 remains dormant, it produces latency associated transcript RNA, or LAT RNA. By using mice, the research team demonstrated that the LAT RNA, extremely unstable, broke down into smaller strands called microRNAs. The microRNAs impede viral reproduction by blocking the synthesis of proteins that aid in the viral replication cycle. Therefore, the virus is able to remain dormant if there are enough microRNAs to suppress protein production. However, when a stressor is present, the virus produces messenger RNA at an amount that the microRNAs cannot block. Thus, the proteins are created and active viral replication proceeds.

Curing the occurrence of cold sores requires a combination therapy. Treatment cannot reach the virus while it is inactive, so it must be activated to kill it. The researchers have designed a drug that binds to the microRNAs responsible for keeping the virus dormant. If the drug works like they expect, it will activate the virus, which would then replicate. After activating the virus, the patient would take acyclovir, which is an antiviral medication that destroys replicating HSV1. This would result in the person being cured of cold sores. Animal trials are being used to determine how the drug can be most effectively delivered. 


It has been years since this discovery, but as we know science takes time. I will definitely keep up with the progression of this research. I hope one day we will find out that no one will ever have to deal with a cold sore again.

References:

http://www.webmd.com/skin-problems-and-treatments/tc/cold-sores-topic-overview#1

https://www.sciencedaily.com/releases/2008/07/080702132213.htm

http://esciencenews.com/articles/2008/07/02/discovery.explains.how.cold.sore.virus.hides.during.inactive.phase

https://www.herpes-coldsores.com/cold_sores.htm

A New Therapy to Combat Drug-Resistant Bacteria

Antibiotics, also referred to as antimicrobial drugs, are one of the most frequently used drugs in medicine. They treat bacterial infections by either killing the bacteria or preventing the bacteria from reproducing and spreading. The emergence of drug-resistant bacteria has reduced the effectiveness of antibiotics. It is projected that approximately 10 million people will die by the year 2050 due to drug-resistant bacteria. Therefore, it is extremely important that new methods are produced to successfully treat bacteria that has adapted over time to resist antibiotics. The researchers suggest that antimicrobial peptides (AMPs) could be a good substitute for current antibiotics. These peptides are present in all living organisms and help defend against microbial infections. Approximately half of the amino acids present in AMPs are hydrophobic, but can adopt amphipathic structures (molecules containing hydrophobic and hydrophilic regions), which allows them to penetrate cell membranes. Franco et. al generated a synthetic peptide known as clavanin-MO. A marine tunicate antimicrobial peptide was used to derive this synthetic peptide.

            Their experiments, which were performed in vitro and in vivo, showed that clavanin-MO contained antimicrobial and immunomodulatory properties. The peptide was successfully active against all the bacterial strains that were tested. Additionally, its efficacy for killing gram-negative and gram-positive bacteria was investigated, and it produced more successful results than the human cathelicidin AMP-LL37. The effectiveness of the peptide was also compared to two commonly used antibiotics, gentamicin and imipenem. Clavinin-MO proved to be more effective, even for multidrug resistant E. coli 2101123. These results exhibit clavanin-MO’s ability to treat a wide range of infections.

            The researchers also performed studies in vitro and in vivo to assess the toxic effects of clavanin-MO. Mouse red blood cells (mRBCs), RAW264.7 macrophage, L929 mouse fibroblast cell lines, and human embryonic kidney cells 293 (HEK-293) were used to perform the experiments. Concentrations of clavanin-MO necessary to produce antimicrobial activity were introduced to RAW264.7, L929 cells, and HEK-293 cells, and no cytotoxic effects were observed. A dose five times the amount necessary for successful antimicrobial treatment was administered to the mice, and still no toxicity was observed. Side effects must be considered when creating new treatments. It is very encouraging that such a large dose did not produce toxic effects. This demonstrates that the treatment can be not only effective, but also safe, which is just as important.

            Clavanin-MO was also found to contain beneficial immunomodulatory properties. One experiment was performed in vivo using the mice to track the migration of leukocytes to the infected area. A major component of treating infections involves inducing leukocytes to travel to the area of infection. Some peptides are able to perform this immunomodulatory activity. The results showed that clavanin-MO significantly increased the number of leukocytes present in the infected area of the mice.

            The increase in antibiotic-resistant bacteria coupled with the lack of antimicrobial therapy development has caused an urgency to create alternative treatments for bacterial infections. Franco et. al have introduced a possible alternative that is very promising. A single peptide, clavanin-MO, has the ability to kill bacteria directly, but can also stimulate immune cells to attack the host. Additionally, throughout the treatment process, the peptide failed to produce toxic effects. While more research is needed, this new therapy moves us considerably closer to a method that combats the rise of antibiotic-resistant bacteria.

References:

http://www.nature.com/articles/srep35465

http://www.cdc.gov/drugresistance/index.html

MutChromSeq: A New Technique for Fast Isolation of Genes

           With the ever-growing human population, it is important to produce enough food to meet the demand. The main source of food production is crops; therefore, it is vital that plants have high yields. Wheat and barley are two major crops found worldwide because they can survive in various environments, result in high yields, and contain many nutrients. If these traits could be controlled, then the yields would increase and there would be food security on a global scale. To control these traits, we must know the genes responsible for them. All we need to do is locate these genes in wheat and barley. This is easier said than done. Wheat and barley have large genomes and large regions of suppressed recombination. This makes it very time-consuming and expensive to use traditional map-based gene isolation. Additionally, obtaining sequences of whole genomes from several mutants is not feasible because comparing large datasets is challenging and costly. There are multiple techniques that can be used to make the sequences of the wheat and barley genomes less complex. However, these methods often result in some sequences being overlooked. Wulff et. al developed a new technique to reduce the complexity of the wheat and barley genomes that does not overlook certain sequences.

Their technique, MutChromSeq, involves flow sorting and comparing the sequences of various mutant chromosomes. This method identifies the mutation and eliminates the need for fine mapping and recombination. MutChromSeq was used to reclone the barley Eceriferum-q gene and clone de novo the wheat Pm2 gene. Even though a false positive was included in the Eceriferum-q example, the correct gene was still identified. The chromosomes were isolated using fluorescent markers. The sequences of the six mutant chromosomes were compared to the wild-type chromosomes.

This approach includes mutagenesis, the reduction of genome complexity, and high-throughput screening. Therefore, the plant species must be able to undergo mutagenesis. Also, the phenotype of the target gene and the chromosome the gene is on must be known. MutChromSeq can be used on genes that meet these requirements, and the method is quick and inexpensive. This method makes it possible to isolate and clone genes that were very difficult to manage in the past.

The researchers introduce a very powerful technique for gene isolation. For genes that could not be easily cloned before, there is now a fast and inexpensive technique that can be used. This is especially impactful for the production of crops. With this new approach, the genes that produce beneficial traits in crops can be targeted. Thus, crops can be improved to better withstand pests and changes in weather. This is very encouraging for the future of food production.

Reference:

Javier Sánchez-Martín, Burkhard Steuernagel, Sreya Ghosh, Gerhard Herren, Severine Hurni, Nikolai Adamski, Jan Vrána, Marie Kubaláková, Simon G. Krattinger, Thomas Wicker, Jaroslav Doležel, Beat Keller, Brande B. H. Wulff. “Rapid gene isolation in barley and wheat by mutant chromosome sequencing.” Genome Biology, 2016.

http://genomebiology.biomedcentral.com/articles/10.1186/s13059-016-1082-1

The Value of Plants in Solving Crimes

            A major aspect of crime scene investigation is the collection of evidence. Plants are likely not the first thing to come to mind when considering valuable evidence for solving a murder. One probably thinks of other types of evidence, such as blood, fingerprints, weapons, or shoe prints. Those are examples of more commonly encountered evidence. However, forensic botany can also provide significant information about a crime that was committed. The anatomy, taxonomy, and ecology of plants are some features of botanical evidence that can be analyzed. The identification of plant species can link the suspect to the crime scene or indicate if the body was moved from one location to another. Additionally, the time a body was buried or left at a site can potentially be determined using plant material.

            Forensic botany involves the use of plants to aid criminal investigations. In 1992, plant DNA was used to solve a murder investigation in Arizona. It was the first time that plant DNA typing was legally accepted as evidence. The body of a woman was discovered in the Arizona desert underneath a Palo Verde tree. A suspect, Mark Bogan, was identified using a beeper found near the body. In the back of his truck, there were seed pods from a Palo Verde tree. Could DNA analysis match the seed pods in the back of his truck to the tree the body was found under? Dr. T. Helentjaris set out to find the answer. A technique called Randomly Amplified Polymorphic DNA (RAPD) was used to analyze the evidence and a band pattern was produced. He also analyzed seed pods collected from other Palo Verde trees throughout Maricopa County to find out if the band patterns of the seed pods were specific to individual Palo Verde trees. He concluded that the seed pods collected from Bogan’s truck came from the tree at the crime scene. He testified in court, and Bogan was convicted of first degree murder.

            Often, botanical trace evidence does not have enough features, especially when the plant material is deteriorated, to identify the genus or species of the plant, so DNA technology is used. Dr. Robert Bever and the Bode Technology Group, Inc. are working on creating molecular methods for analyzing botanical trace evidence, which can be very useful for linking evidence and/or suspects to the crime scene. Using plant systematics, many loci that aid in the identification of plants have been characterized, which include several nuclear (18S, ITS1, ITS2) and chloroplast (rbcL, atpB, ndhF) genes. A DNA extraction, cloning, and sequencing procedure for identifying plants has been created by the Bode Technology Group, Inc. using some of those genes. Applying these methods, many plant species have been identified from physical evidence, such as algae, evergreens, and flowering herbs, shrubs, and trees. Some plants are located in very specific areas in which botanical trace evidence could be extremely valuable for establishing linkage.

            I provided one example of how plant material can be used as forensic evidence, but the attached article contains many more cases that exemplify the impact botanical evidence can have in solving crimes. Many of these investigations would likely not have been solved without the collection of plant materials as evidence. This emphasizes the importance of considering all types of items and materials as evidence because the seemingly smallest piece of evidence could be the key to solving the crime. Hopefully more attention will be given to this type of evidence when analyzing crime scenes given the impact it could have on the investigation.

References:

http://www.cmj.hr/2001/42/3/11387649.pdf

http://www.denverda.org/DNA_Documents/bogan.pdf

A Combined Therapy Technique for Treating Cancer

Many people are diagnosed with cancer every day or have loved ones who have suffered from all different types of cancer. With so many people battling cancer, effective treatment strategies are crucial. Chemotherapy is one type of current cancer treatment that is widely used. However, it has its drawbacks. Patients experience many harmful side effects because the drugs target both cancerous and healthy cells. Photothermal therapy (PTT) is an emerging technique that has shown to be effective in killing cancer cells. It uses electromagnetic energy (usually in near infrared wavelength) to excite light sensitizers (such as noble metals, carbon materials, or organic polymers and molecules). When excited, they release vibrational energy in the form of heat, which kills the cancer cells. This technique is especially favorable because it is selective, targeting the cancer cells and leaving healthy cells alone. Therefore, it is noninvasive and much safer. Adverse side effects are severely diminished. However, PTT does not always destroy all the cancer cells, which can result in relapses.

            The limitations of these treatments lead researchers to consider a combined treatment therapy that involves chemotherapy and PTT. Studies have shown that heat can improve the effectiveness of chemotherapy, so researchers have focused on the cancer treatments that combine chemotherapy and PTT. Tu et. al developed carbon nanoparticles (CNPs)/doxorubicin@SiO₂ nanocomposites via reverse microemulsion, while also controlling the size and high drug loading ratio, to deliver the drugs using the combined methodology. The nanocomposites are a good platform for delivery due to their ability to generate high heat, perform pH responsive drug delivery, and release a high amount of drugs when induced by heat. Mice were injected with 4T1 tumor cells. They were treated using each therapy separately and combined. For the single therapy, the tumors continually grew throughout 14 days of treatment. When the treatment techniques were combined, the tumors were completely eradicated six days after the treatment. These results exhibit the effectiveness of a combined therapy approach for cancer treatment.

Figure 1. Schematic diagram showing the strategy for the synthesis and combined photothermal and chemo-therapy of CNPs/Dox@SiO₂ NCs.

This research is very important given the prevalence of cancer throughout the world. In the United States alone, approximately 1,685,210 cancer cases will be diagnosed and 595,690 deaths from cancer will occur in 2016 (American Cancer Society). With the large incidence of cancer diagnoses and cancer deaths, the demand for newer, safer, and more effective treatments is great. This study introduces an alternative technique for cancer treatment that combines chemotherapy and photothermal therapy with the use of carbon nanoparticles (CNPs)/doxorubicin@SiO₂ nanocomposites to deliver the drugs. This is a more effective technique due to the selective targeting of cancer cites. This therapy would produce significantly fewer side effects for patients. Additionally, the method’s ability to eradicate the entire tumor severely decreases the chances that the patient will have a recurrence of the cancer. This methodology also has such a huge impact because it can be used on all types of cancer.

References:

http://www.cancer.org/research/cancerfactsstatistics/index

Xiaolong Tu, Lina Wang, Yuhua Cao, Yufei Ma, He Shen, Mengxin Zhang, and Zhijun Zhang. "Efficient cancer ablation by combined photothermal and enhanced chemo-therapy based on carbon nanoparticles/doxorubicin@SiO2 nanocomposites." Carbon. Volume 97, 35-44, February 2016. http://www.sciencedirect.com.ezproxy.shsu.edu/science/article/pii/S0008622315004406

Does an apple a day really keep the doctor away?

            I love apples, and I eat one every day with my lunch. The saying “an apple a day keeps the doctor away” often comes to mind. This lead me to think: is it actually true? So I did some research and this is what I found.

            In 2000, researchers at Cornell University found that eating an apple a day does help keep the doctor away, publishing their work in the scientific journal, Nature. They found that phytochemicals, mostly found in apple’s skin, provide anticancer and antioxidant benefits. They also claim that eating whole fruits and vegetables provides more health benefits than vitamin supplements. Cornell’s Food Science and Toxicology department studies found that the antioxidant properties in a fresh apple were equal to 1,500 milligrams of Vitamin C.

            Liu et. al looked at the difference in effects on cancer cell growth between the apple’s skin and its flesh. Colon cancer cells were treated with apple extract from the skin and flesh of a Red Delicious apple. The cancer cell growth was decreased by 43 % using 50 milligrams of the apple skin extract, while only 29 % of the cancer cell growth decreased when using the apple extract without the skin. The extracts were also tested on liver cancer cell growth. Similar results were obtained, with the apple extract from the skin and flesh decreasing cancer cell growth by 57 % and 40 %, respectively.

A more recent article on the topic was published in the Journal of Functional Foods in 2013. Zhao et. al discussed how apple consumption decreased plasma concentrations of oxidized low-density lipoprotein/beta₂-glycoprotein I complex (oxLDL-β₂GPI), which is a contributor to atherosclerosis. Atherosclerosis is a condition characterized by fatty plaques in the arteries, which can lead to cardiac events such as heart attack and stroke.

These studies show that apples can indeed help keep the doctor away!

References:

http://www.webmd.com/food-recipes/news/20000621/benefits-of-eating-fruit#1

Eberhardt, M. V., Lee, C. Y., and Liu, R. H. Antioxidant activity of fresh apples. Nature 405(6789): 903-904, 2000.

Link to abstract: http://www.nature.com/nature/journal/v405/n6789/full/405903b0.html

Shi Zhao, Joshua Bomser, Elizabeth L. Joseph, and Robert A. DiSilvestro. Intakes of apples or apple polyphenols decease plasma values for oxidized low-density lipoprotein/beta₂-glycoprotein I complex. Journal of Functional Foods. Volume 5, Issue 1, 493-497, January 2013.

http://www.sciencedirect.com.ezproxy.shsu.edu/science/article/pii/S1756464612001363