Nanodevices built with DNA origami

Did you know DNA could be used for origami?

Not actual DNA origami.

Not actual DNA origami.

The precise control and organization of nanoscale devices has shown a great potential for ultimately creating “nano-devices” that can perform nanoscale biological measurements, deliver medicine in vivo, among many other applications. A recent article from Carlos E. Castro and colleauges from The Ohio State University demonstrates the use of DNA origami with programmable complex and reversible 1D, 2D and 3D motions.

By varying the DNA origami design, they were able to observe different mechanisms for the DNA origami’s 3D motion such as the crank-slider and four bar mechanism. The research team mainly utilized transmission electron microscopy (TEM) to follow the morphology changes as the origami moves.

DNAUsing a fluorescence quenching assay (attaching a fluorescent label on one arm and a quencher on the other), they have characterized the timescale of DNA origami motion. Overall, their group sees this technology as a “foundation for developing and characterizing a library of tunable DNA origami kinematic joints and using them in more complex controllable mechanisms similar to macroscopic machines, such as manipulators to control chemical reactions, transport biomolecules, or assemble nanoscale components in real time.”

 

Shown below are some of the videos showing the motions of the DNA origami that they have reported:

About the author: Herdeline Ann M. Ardoña is a third year graduate student at Johns Hopkins University Department of Chemistry, currently working in chemistry professor J.D. Tovar’s lab and co-advised by professor Hai-Quan Mao, in materials science and engineering.

Reference: Programmable motion of DNA origami mechanisms. (Proc. Natl. Acad. Sci. U.S.A., 2015, 112, 713-718)

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, mspiro@jhu.edu or 410-516-4802.

Prizes offered for top poster presenters

We need your posters! INBT’s annual symposium theme relates to neuroscience, but posters on any multidisciplinary topic are encouraged. Submission deadline for posters is April 27. Posters will be judged and prizes will be awarded to top presenters!

Erlenmeyer_Flasks.-awardJohns Hopkins Institute for NanoBioTechnology hosts its annual symposium May 1 in the Owens Auditorium (between CRB I and CRB II) at the medical campus. Faculty expert speakers present in the morning on our theme, Neuro X, where x can be medicine, engineering, science, etc. The poster session begins in the afternoon. Posters on ANY MULTIDISCIPLINARY TOPIC are encouraged, and we welcome submissions from any department or division. Prizes will be awarded to top presenters. Submission guidelines, the full speaker agenda and additional information can be found online. Submit your poster now at http://inbt.jhu.edu/news/symposium/

Are there problems with the peer-review process?

The pathway to publication is littered with checkpoints, reviews, and rejections. Before your paper is accepted it is read and reviewed with a few possible fates. It can be desk rejected by the editor and never reviewed or it can reach the reviewers who then decide the fate of the manuscript.

questionamarkwebSiler et al. investigated the effectiveness of the review process. They observed that top ranking journals overall have a very effective desk screening process where the best manuscripts are selected for review. However, there was one main fault; these top tier journals desk rejected the top cited manuscripts. This is likely due to the fact that their goal is to publish papers that are widely applicable and of interest to many people. This limits the ability of truly novel and exciting works to be published in these formats.

Overall, however, it was determined that the review process is helpful. Manuscripts that went to review overall had more citations than those desk rejected and resubmitted elsewhere. The results of this study were reassuring, and it was nice to see that at least a few scientists are looking into the effectiveness of the review process.

Link to article: http://www.pnas.org/content/112/2/360.abstract

About the author: Moriah Knight is a third year in the Johns Hopkins Department of Materials Science and Engineering working in Peter Searson’s lab.

Hanes to be keynote speaker for student symposium

Justin Hanes, PhD, director of the Center for Nanomedicine at the Johns Hopkins School of Medicine and affiliated faculty member of the Institute for NanoBioTechnology, will be the keynote speaker of the student-organized mini symposium held at 9 a.m., March 24 in The Great Hall at Levering on the Homewood campus. Student presenters affiliated with INBT, Center for Cancer Nanotechnology Excellence and Physical Sciences-Oncology Center laboratories will round out the rest of the program.  This event is free and open to the entire JHU community. Refreshments provided.

INBT Mini Symposium 2015 Poster 2 (1)Agenda

  • 9:00 – 9:30 am: Light Breakfast & Introduction
  • 9:30 – 9:50 am: Alyssa Kosmides, Biomedical Engineering
    Nano-scale platforms for tumor specific CD8 cell activation & manipulation
  • 9:50 – 10:10 am: Quinton Smith, Chemical & Biomoleular Engineering
    Determining the physiochemical cues directing stem cell fate
  • 10:10 – 10:30 am: Lye Lin Lock, Chemical & Biomolecular Enigneering
    Protease-activatable supramolecular nanobeacon for cancer diagnostic and therapeutic applications
  • 10:30 – 10:40 am: Break
  • 10:40 – 11:00 am: Alexander Komin, Materials Science & Engineering
    Investigating the permeation of cell monolayers by cell penetrating peptides
  • 11:00 – 11:20 am: Daniel Shea, Chemical & Biomoleular Engineering
    Kinetic properties govern Mucin 16 & Podocalyxin adhesion to E-& L-selectins in shear flow
  • 11:20 – 11:30 am: Break
  • 11:30 – 12:30 pm:  Justin Hanes, PhD, the Lewis J. Ort Professor of Ophthalmology
    Director of The Center For Nanomedicine at the Wilmer Eye Institute

Reception to follow.

Nano-bio lab course: MAPs and CD

Editor’s note: Over the next several days, we will share the student impressions of some of the techniques learned in INBT’s nano-bio laboratory course (670.621). These reports demonstrate the wide variety of techniques students trained at the Johns Hopkins Institute for NanoBioTechnology are expected to understand. Each technique is taught in a different affiliated faculty lab. More lab techniques to come.

Membrane Active Proteins (MAPs) and Circular Dichroism (CD) spectrography

During this lab, we learned a couple of techniques that I had not used before. First we synthesized liposomes and processed them, resulting in uniform liposome radius. Then we made a solution of membrane active proteins with aromatic amino acids so that their absorbance and emission could be measured.

CD_trans

circular dichroism (CD) spectrography

We ran the proteins through the fluorometer at varying wavelengths to create a profile of emission and absorbance of the protein. This was done also at varying pHs and at different liposome concentrations.

In theory the proteins should incorporate into the liposomes and there should be a change in the spectra as a result. During our lab time we had issues getting the desired results, but it was still informative on how to use the fluorometer and other new equipment. We found the spectra for two different proteins at two different pH values for each to see the effect that pH had on the emission/absorbance spectra.

We also preformed CD spectrography (circular dichroism) to determine the chirality of the proteins, that is, how are the specific molecules spatially arranged. Again the procedure did not work exactly as planned, but learning how to perform the measurement was informative, nonetheless.

About the author: Jackson DeStefano is a first year PhD candidate in the laboratory of Peter Searson, professor of materials science and engineering.

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, mspiro@jhu.edu or 410-516-4802.
.

Nanoparticle-based medicine prevents rejection in corneal implants

Of the 48,000 corneal transplants performed each year in the United States, 10 percent result in rejection because of poor medicine compliance. A biodegradable nanomedicine developed by researches affiliated with the Johns Hopkins School of Medicine and the Institute for NanoBioTechnology (INBT) could provide the controlled-release medications needed after eye surgery that improve corneal transplant outcomes.

“Medicine compliance is a major challenge in patient care,” says Walter Stark, M.D., chief of the Division of Cornea, Cataract and External Eye Diseases at Johns Hopkins. “About 60 to 80 percent of patients don’t take medicine the way they are supposed to.”

In an animal study being published in the March 10 issue of the Journal of Controlled Release, researchers looked into ways to alleviate the strain of adhering to a post-surgery treatment regimen that is sometimes hard to manage.

Corneal transplant

Corneal transplant

Rats that underwent a corneal graft surgery were randomly divided into four groups and were given various treatments. One group was injected weekly for nine weeks with a safe, biodegradable nanoparticle loaded with corticosteroids for timed release of medicine. The other three groups received weekly injections of saline, placebo nanoparticles and free dexamethasone sodium phosphate aqueous solution after surgery, respectively.

Treatments were given until the graft was clinically deemed as failed or until the nine-week test period concluded. Researchers looked at corneal transparency, swelling and growth of new blood vessels to decide if a graft had failed. For rats that received the nanoparticle loaded with corticosteroids, 65 percent of the treatment remained in the eye and did not leak within one week of the surgery. The concentration of the treatment also remained stronger than in the other three treatment groups. Additionally, there were no signs of swelling, and the cornea was clear throughout the test period. There were also far fewer instances of unwanted growth of new blood vessels in this group.

Two weeks after surgery, rats that received the placebo nanoparticle and saline injections had severe swelling, opaque corneas and unwanted growth of new blood vessels, all indicating graft failure. After four weeks, rats that received free dexamethasone sodium phosphate aqueous solution all had graft failure as well. The only group that showed successful corneal transplant was the group of rats that received the corticosteroid-loaded nanoparticle injections. The grafts were still viable in 100 percent of these rats.

“Corneal grafts are not easy to come by, and a lot of testing and time goes into ensuring the safe use of a graft for cornea transplant,” says Qingguo Xu, Ph.D., a research associate at the Center for Nanomedicine at the Wilmer Eye Institute at Johns Hopkins Medicine. “This is why we want to do a better job at making sure corneal transplants don’t end up in rejection, and our study illustrates a potentially better way.”

The steroid-loaded nanoparticle treatment group showed no signs of corneal transplant rejection. “That’s 100 percent efficacy, a very promising finding,” says Justin Hanes, Ph.D., director of the Center for Nanomedicine and INBT affiliate. “This type of treatment may also help prevent corneal transplant rejection in humans while making medicine adherence much easier on patients and their families.”

The nanoparticle loaded with medication could eliminate the need for a patient to remember to take their medicine ― often multiple doses per hour ― after a surgery, alleviating compliance risk. These types of drug delivery systems could be paired with other drugs and used in other conditions, such as glaucoma, macular degeneration and corneal ulcers, among others. The research team intends to continue the collaboration between engineering and medicine to look further into better ways to treat eye diseases.

Additional authors include Qing Pan, Nicholas J. Boylan, Nicholas W. Lamb, David Emmert, Jeh-Chang Yang, Li Tang, Tom Feflin, Saeed Alwadani and Charles G Eberhart.

Funding of this study came from the Raymond Kwok Family Research Fund. This work was also partially funded by a grant from the King Khaled Eye Specialist Hospital of Saudi Arabia and the Eye Bank Association of America/Richard Lindstrom Research Grant 2012.

Note: Justin Hanes will be the keynote speaker for INBT’s student symposium to be held March 24 in the Great Hall at Levering on the Hopkins Homewood campus. To RSVP to the student symposium, visit our Facebook event page here: https://www.facebook.com/events/648159565330499/

For more Johns Hopkins Medicine news go to http://www.hopkinsmedicine.org/news/media/releases

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, mspiro@jhu.edu or 410-516-4802.

How firefly research helped gene therapy

Sometimes on a calm summer or fall night, one is able to observe the beautiful dance of blinking fireflies. Scientists began to explore mechanisms to describe this unique natural phenomenon as early as the late 1800’s. After a series of experiments with solutions at different temperature with ground up abdomens of fireflies, Raphael Dubois named the enzyme luciferase and the substrate luciferin that were the cause of the light-producing reaction (1).  But it wasn’t until recently in 1985 that scientists were able to clone the gene for luciferase and express it in bacteria to produce the luciferase.

firefly

Figure 1: Picture of firefly. Source: http://www.fireflyexperience.org/photos/

Once the gene was cloned, genetic researchers realized the importance of the findings and started to use it as a reporter gene for experimental gene therapy. Gene therapies involve transfection of new genetic material into the host’s DNA and can be applied not only for therapies for diseases of genetic origin, but can be used for cancer therapy and diagnostic purposes.

By incorporating the gene for luciferase along with the gene of interest, the Hai-Quan Mao lab in the Department of Materials Science and Engineering at Johns Hopkins University can detect whether or not their nanoparticles used for gene delivery have been successful simply by adding luciferin to the cells. If the gene transfer was successful, then the luciferase will act on the substrate luciferin to emit light.

Sources

1)     Fraga, Hugo. “Firefly luminescence: A historical perspective and recent developments.” Photochemical & Photobiological Sciences 7.2 (2008): 146-158.

About the author: John Hickey is a second year Biomedical Engineering PhD candidate in the Jon Schneck lab researching the use of different biomaterials for immunotherapies and microfluidics in identifying rare immune cells.

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, mspiro@jhu.edu or 410-516-4802.

 

Image and video contest celebrates 15 years of nano

In honor of the 15th Anniversary of President Clinton’s landmark speech announcing the National Nanotechnology Initiative, the NNI is hosting a contest for video and photography. Submit your photos! Submit your videos!

Screen Shot 2015-03-09 at 5.13.14 PMVideo contest

Will your research lead to nanotechnologies that impact our daily lives? Demonstrate how your work will bring solutions to real‐world problems. This contest is for graduate students in the United States and U.S. territories.

Video submissions are accepted from January 21, 2015 through April 24, 2015. Public voting begins begins May 1, 2015.
For details visit: http://www.nano.gov/studentvideocontest

Questions? Email us: nncovideos@gmail.com

Image contest

Help us demonstrate how beautiful the nanoscale can be and explain how the research behind your picture may lead to nanotechnologies that benefit society. This contest is for students in the United States and U.S.territories.

Photographs accepted from January 20, 2015 through March 31, 2015. Public voting begins April 7, 2015.
For details visit: http://www.nano.gov/envisionano
Questions? Email us: envisionano@nnco.nano.gov

Brought to you by the National Nanotechnology Initiative: www.nano.gov

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, mspiro@jhu.edu or 410-516-4802.

 

Nano-bio lab course: micro- printing and patterning

Editor’s note: Over the next several days, we will share the student impressions of some of the techniques learned in INBT’s nano-bio laboratory course (670.621). These reports demonstrate the wide variety of techniques students trained at the Johns Hopkins Institute for NanoBioTechnology are expected to understand. Each technique is taught in a different affiliated faculty lab. More lab techniques to come.

Micro- printing and patterning

In this lab we learned the techniques associated with microcontact printing. A patterned wafer, like the ones created last week in photolithography, was used to create a PDMS stamp (PDMS is a type of silicone) with specific surface features according to our design. This stamp was then coated in proteins and used to transfer the pattern to a plasma cleaned glass microscope slide.

micropatternThis technique is of interest to me because it has the possibility to be incorporated into microfluidic devices. In our experiments we normally coat channels with fibronectin or collagen in order to increase cell adhesion to the surface. With the use of microcontact printing it may be possible to lay down specific proteins in much more precise locations in order to study the behavior of cells.

One drawback that may arise from trying to use microcontact printing in conjunction with a microfluidic device would be aligning the pattern of protein that was laid down with the pattern of the microfluidic device. Normally the PDMS of a device is plasma-bonded by hand to the glass slide, thus there is only as much precision as the eye. This would mean that the device would have to be general enough so that you could be sure you don’t overlap the patterns.

About the author: Jackson DeStefano is a first year PhD candidate in the laboratory of Peter Searson, professor of materials science and engineering.

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, mspiro@jhu.edu or 410-516-4802.

O-GlcNAc: The Sweet Side of Epigenetics

In 1992 Edmond H. Fischer and Edwin G. Krebs won the Nobel Prize in Physiology or Medicine “for their discoveries concerning reversible protein phoshorylation as a biological regulatory mechanism.” Phosphorylation of proteins can essentially be thought of as the on/off switch that regulates protein activity inside of cells.

It became increasingly clear later on, however, that protein physiology was much more complex than regulation through just a simple on/off phosphorylation switch. It was eventually discovered by Johns Hopkins’ very own Dr. Gerald Hart that a very special sugar called N-acetylglucosamine (GlcNAc) can be added to the same places on proteins where phosphorylation often occurs. The addition of GlcNAc to these sites is now known as the O-GlcNAc modification. O-GlcNAc essentially serves as another layer of control over protein physiology by acting as a sort of “cap” that must be removed before a protein can be phosphorylated. In otherwords, phosphorylation and the O-GlcNAc modification cycle between each other to regulate how many important proteins behave. One amazing feature of the O-GlcNAc modification is the fact that it is performed by only two enzymes, OGT which adds it to proteins and OGA which removes it, and that’s it. This is in stark contrast to protein phosphorylation and dephosphorylation which needs hundreds of different enzymes to perform phosphorylation mechanics.

Fig 1.  Histones are modified by O-GlcNAc.

Fig 1. Histones are modified by O-GlcNAc.

To this day O-GlcNAc cycling remains an enigma, however, emerging evidence continues to mount that illustrates the very important physiological roles for O-GlcNAc. Two of some of the most important concepts within the realm of epigenetics are the modifications of histones and the methylation of DNA. It is now known that histones, which are proteins that help package DNA into the nucleus, are modified by O-GlcNAc 1 (fig 1.). The other major type of epigenetic regulation of gene expression– methylation of DNA–silences genes, but is also a reversible process. Proteins named TETs help to remove methyl groups on DNA to reverse this silencing. Recently it has also been shown that TETs have their activity regulated by O-GlcNAc 2. In otherwords, O-GlcNAc seems to have a very important role in regulating and interacting with two very important physiological mechanisms that write the epigenetic code.

Finally, glucose is most often thought of as fuel for the cell–and this is true–however, the substrate that is required to perform the O-GlcNAc modification (GlcNAc) happens to also be a byproduct of glucose metabolism. Major diseases such as cancer, diabetes, and Alzheimer’s are often associated with altered glucose metabolism and also have profound epigenetic changes. It is quite tempting, therefore, to postulate that O-GlcNAc may be the key that links environment, stress, nutrient availability, and metabolism to changes in epigenetics. Understanding carbohydrate metabolism and O-GlcNAc regulation of epigenetics may one day open new doors that will lead to breakthroughs in regenerative medicine, understanding embryological development, tissue engineering, and treating major diseases.

About the Author: Christopher Saeui is a fourth year Biomedical Engineering PhD student in the Kevin J. Yarema Laboratory for Cell and Carbohydrate Engineering studying the epigenetic and metabolic mechanisms that alter glycosylation in cancer.

References
1. Sakabe, K., Wang, Z. & Hart, G. W. Beta-N-acetylglucosamine (O-GlcNAc) is part of the histone code. Proc. Natl. Acad. Sci. U. S. A. 107, 19915-19920 (2010).
2. Shi, F. T. et al. Ten-eleven translocation 1 (Tet1) is regulated by O-linked N-acetylglucosamine transferase (Ogt) for target gene repression in mouse embryonic stem cells. J. Biol. Chem. 288, 20776-20784 (2013).

For all press inquiries regarding INBT, its faculty and programs, contact Mary Spiro, mspiro@jhu.edu or 410-516-4802.