Featured Research

from universities, journals, and other organizations

Pressured Proteins: A Little Pressure In Proteomics Squeezes 4-hour Step Into A Minute

Date:
July 12, 2008
Source:
DOE/Pacific Northwest National Laboratory
Summary:
Many coaches inspire better performance by pressuring their teams. Now, proteomics researchers are using pressure to improve the performance of their analyses. In a simple solution to a time-consuming problem, the researchers have found that adding pressure early in their protocol squeezes four hours of waiting -- often allowed to last overnight for convenence -- into a minute. The result brings researchers closer to "proteomics on the fly."

Many coaches inspire better performance by pressuring their teams. Now, proteomics researchers are using pressure to improve the performance of their analyses. In a simple solution to a time-consuming problem, the researchers have found that adding pressure early in their protocol squeezes four hours of waiting into a minute.

Related Articles


"We were really happy to see how well it worked," said biochemist Daniel Lopez-Ferrer, a post-doctoral researcher at the Department of Energy's Pacific Northwest National Laboratory. "We're determining when and how to incorporate it into our analyses."

Proteomics researchers learn about organisms by studying the proteins they make, and they want to be able to analyze large numbers of samples fast and sensitively. Cutting down the time-consuming steps would significantly increase the number of samples that could be analyzed. The first, slowest steps in most proteomics protocols requires at least four hours -- researchers often allow this to go overnight because of timing issues for the rest of the process. The team of chemists, biologists and physicists at PNNL investigated whether pressure would improve their process.

What takes so long is the breakdown of full-length proteins into smaller bits called peptides, which can be analyzed by measuring their individual masses. The most common way to break down proteins is to let an enzyme eat through the bonds between the protein building blocks, or amino acids. Scientists have tried a variety of ways to help the enzyme digest these bonds faster, including using microwaves or ultrasound, with varying degrees of success.

Food processors have long been using high pressure to kill pathogens on food (think homemade preserves), and some evidence suggested that as pressure rises from low to high, some enzymes initially become more active before dying from the duress.

To determine if pressure would help in proteomics, the team used the protein albumin. The researchers incubated albumin with the enzyme trypsin at several different pressures for one minute each, then counted how many pieces into which trypsin cut albumin. At 10,000 pounds-per-square-inch up to 35,000 psi, trypsin appeared to maximally cleave albumin. For these tests, the team used facilities at the DOE's Environmental Molecular Sciences Laboratory on the PNNL campus.

The team then tested whether this process could be used for proteomics, in which large numbers of proteins are studied at once. So, they extracted the proteins from a bacterial culture and subjected half to the traditional overnight approach and half to 35 kpsi for one minute.

Overall, the resulting collection of peptides looked very similar between the two methods. The pressure method generated about 10 percent more unique peptides but cut slightly fewer total bonds than the traditional method. The researchers concluded that the benefit of time gained by the pressure method and the additional unique peptides outweighed the dip in total bonds.

Additionally, the team wondered how pressure sped up the enzyme digestion. To determine if pressure caused bacterial proteins to unfold and make their bonds more vulnerable to trypsin, the team turned to myoglobin, a compact glob of a protein that packs a second, loose molecule inside. When they put myoglobin under 35 kpsi in the absence of trypsin, myoglobin lost its molecular parcel, suggesting that pressure opens up or denatures proteins, baring their bonds and giving trypsin more room to work.

Now, the PNNL researchers are integrating the pressurized digestion into their proteomics protocol where appropriate. In other work, Lopez-Ferrer and colleagues are testing whether ultrasound would also speed up this first step. They believe pressure and ultrasound might bring different advantages to the table of protein digestion, depending on the samples to be analyzed.

This work was initially supported by internal PNNL funds, and was then funded by the National Institutes of Health.


Story Source:

The above story is based on materials provided by DOE/Pacific Northwest National Laboratory. Note: Materials may be edited for content and length.


Journal Reference:

  1. Lo%u0301pez-Ferrer et al. Application of Pressurized Solvents for Ultrafast Trypsin Hydrolysis in Proteomics: Proteomics on the Fly. Journal of Proteome Research, 2008; 0 (0): 0 DOI: 10.1021/pr7008077

Cite This Page:

DOE/Pacific Northwest National Laboratory. "Pressured Proteins: A Little Pressure In Proteomics Squeezes 4-hour Step Into A Minute." ScienceDaily. ScienceDaily, 12 July 2008. <www.sciencedaily.com/releases/2008/07/080709144132.htm>.
DOE/Pacific Northwest National Laboratory. (2008, July 12). Pressured Proteins: A Little Pressure In Proteomics Squeezes 4-hour Step Into A Minute. ScienceDaily. Retrieved October 31, 2014 from www.sciencedaily.com/releases/2008/07/080709144132.htm
DOE/Pacific Northwest National Laboratory. "Pressured Proteins: A Little Pressure In Proteomics Squeezes 4-hour Step Into A Minute." ScienceDaily. www.sciencedaily.com/releases/2008/07/080709144132.htm (accessed October 31, 2014).

Share This



More Plants & Animals News

Friday, October 31, 2014

Featured Research

from universities, journals, and other organizations


Featured Videos

from AP, Reuters, AFP, and other news services

Watch Baby Goose Survive A 400-Foot Cliff Dive

Watch Baby Goose Survive A 400-Foot Cliff Dive

Buzz60 (Oct. 31, 2014) For its nature series Life Story, the BBC profiled the barnacle goose, whose chicks must make a daredevil 400-foot cliff dive from their nests to find food. Jen Markham has the astonishing video. Video provided by Buzz60
Powered by NewsLook.com
World's Salamanders At Risk From Flesh-Eating Fungus

World's Salamanders At Risk From Flesh-Eating Fungus

Newsy (Oct. 31, 2014) The import of salamanders around the globe is thought to be contributing to the spread of a deadly fungus. Video provided by Newsy
Powered by NewsLook.com
Alcoholic Drinks In The E.U. Could Get Calorie Labels

Alcoholic Drinks In The E.U. Could Get Calorie Labels

Newsy (Oct. 31, 2014) A health group in the United Kingdom has called for mandatory calorie labels on alcoholic beverages in the European Union. Video provided by Newsy
Powered by NewsLook.com
Malaria Threat in Liberia as Fight Against Ebola Rages

Malaria Threat in Liberia as Fight Against Ebola Rages

AFP (Oct. 31, 2014) Focus on treating the Ebola epidemic in Liberia means that treatment for malaria, itself a killer, is hard to come by. MSF are now undertaking the mass distribution of antimalarials in Monrovia. Duration: 00:38 Video provided by AFP
Powered by NewsLook.com

Search ScienceDaily

Number of stories in archives: 140,361

Find with keyword(s):
Enter a keyword or phrase to search ScienceDaily for related topics and research stories.

Save/Print:
Share:

Breaking News:

Strange & Offbeat Stories


Plants & Animals

Earth & Climate

Fossils & Ruins

In Other News

... from NewsDaily.com

Science News

Health News

Environment News

Technology News



Save/Print:
Share:

Free Subscriptions


Get the latest science news with ScienceDaily's free email newsletters, updated daily and weekly. Or view hourly updated newsfeeds in your RSS reader:

Get Social & Mobile


Keep up to date with the latest news from ScienceDaily via social networks and mobile apps:

Have Feedback?


Tell us what you think of ScienceDaily -- we welcome both positive and negative comments. Have any problems using the site? Questions?
Mobile: iPhone Android Web
Follow: Facebook Twitter Google+
Subscribe: RSS Feeds Email Newsletters
Latest Headlines Health & Medicine Mind & Brain Space & Time Matter & Energy Computers & Math Plants & Animals Earth & Climate Fossils & Ruins