Navigation Links
Pressured proteins: A little pressure in proteomics squeezes 4-hour step into a minute
Date:7/9/2008

RICHLAND, Wash. -- 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.

"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." Lopez-Ferrer and his colleagues reported their findings in July 8, 2008 Journal of Proteome Research.

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.


'/>"/>

Contact: Mary Beckman
mary.beckman@pnl.gov
509-375-3688
DOE/Pacific Northwest National Laboratory
Source:Eurekalert

Related medicine news :

1. Hormone Therapy Shows Little Benefit Against Prostate Cancer
2. Blue Cross Blue Shield of Michigans Proposed Reforms Make Little Progress With State Senate
3. Homeopathic First Aid Kit: Solutions for Summers Little Bummers
4. So Much to Do, So Little Sleep
5. Societys attitudes have little impact on choice of sexual partner
6. Kroger and The Little Clinic Establish Partnership to Offer Customers Convenient Healthcare Centers Inside Their Neighborhood Store
7. Batman Takes on Gotham City With a Little Help From Milk
8. Dialysis Catheter Placement Makes Little Difference in Infection Risk
9. Corticosteroids of Little Use Against Childhood Meningitis
10. Come to Maggianos Little Italy(R) to Eat-A-Dish for Make-A-Wish(R)
11. New Independent Research Firm Targets Firms with Little or No Analyst Following
Post Your Comments:
*Name:
*Comment:
*Email:
(Date:2/21/2017)... Marne, Michigan (PRWEB) , ... February 21, 2017 ... ... contributions to the United States during Black History Month (officially African American ... additions to many fields of American society of a diverse race of people, ...
(Date:2/21/2017)... ... 21, 2017 , ... CLICKco LLC, a company dedicated to ... now available for purchase on RonnieColemanNutrition.com, a popular website for health and nutritional ... consumers who love coffee but are looking to add more protein to their ...
(Date:2/20/2017)... Boca Raton, FL (PRWEB) , ... February 20, ... ... for creating a patent-pending probiotic supplement that supports the proper levels of “good” ... for purchase on StackedNutrition.com, a popular nutritional products website. , Daily Body ...
(Date:2/20/2017)... (PRWEB) , ... February 20, 2017 , ... ... and check-in, a cloud-based self-service technology that runs on the Posiflex XT-series of ... No. 1076 during HIMSS17 in Orlando, Florida. The terminal provider Posiflex ...
(Date:2/20/2017)... (PRWEB) , ... February 20, 2017 , ... ReportingMD, a Population Health Software Company, is ... of the company. The new location will triple the size of the previous location ... the town of Sunapee, NH. , “We are excited to expand our footprint ...
Breaking Medicine News(10 mins):
(Date:2/21/2017)... 21, 2017 Mass Spectrometer Market: Overview ... This report on mass spectrometer ... prospects of the market globally. The stakeholders of ... the manufacture and commercialization of various mass spectrometer ... planning to enter this market. This report comprises ...
(Date:2/21/2017)... -- Luminex Corporation (NASDAQ: LMNX ) (the "Company") today ... of a quarterly cash dividend to its shareholders, the first ... payable on April 14, 2017 to shareholders of record as ... The board of directors intends for the Company ... holders of its common stock, representing a planned annual dividend ...
(Date:2/21/2017)... Feb. 21, 2017  Global health services company Cigna ... ® for breast reconstruction surgery and Dermacell ... an advanced acellular dermal matrix developed by LifeNet Health ... NVDQ ) "We are proud ... health-care payers to review the growing base of evidentiary ...
Breaking Medicine Technology: