Labs around the world are using Waters’ Synapt HDMS systems to advance drug discovery, biological molecule profiling, and infectious disease research.

In January, Waters Corp., Milford, Mass., launched the Waters Synapt mass spectrometry (MS) system, a next-generation quadrupole orthogonal acceleration (OA) time-of-flight (TOF) mass spectrometry platform. The Synapt MS system provides an upgrade pathway to the Synapt High Definition MS (HDMS) system, which enables researchers to analyze samples differentiated by size, shape, and charge, as well as mass, ultimately providing new capabilities that can help them meet and exceed future requirements.

“Confident sample identification, detailed characterization, and increased productivity are primary requirements for intelligent MS-based solutions in biomedical applications such as proteomics, metabonomic profiling, biomarker discovery/validation, and pharmaceutical R&D,” says Brian Smith, VP of MS operations for the Waters Division.

“The new Synapt MS system meets these demands through application-specific system solutions designed to help our customers accelerate and improve the quality of laboratory analysis with the goal of advancing research and reducing time-to-market.”

The Synapt MS system combines Waters’ Acquity UltraPerformance LC (UPLC) separations, Exact Mass MSE data acquisition, and MassLynx Informatics—designed to generate high quality, comprehensive data from complex biological samples, maximize confidence in results, and enable scientists to make better informed decisions.

“A unique aspect of the Synapt MS system is that it is HDMS-ready,” adds Smith. “Our customers will be able to upgrade their systems to incorporate the HDMS capabilities when they require more powerful solutions to meet their future goals, effectively future-proofing their laboratories.”

Advancing drug discovery

Using Waters’ TriWave technology, Synapt HDMS systems combine high-efficiency, ion mobility-based measurements and separations with high-performance quadrupole TOF mass spectrometry. Synapt HDMS systems enable researchers to differentiate molecules by size, shape, and charge, as well as mass and to deliver increased specificity and sample definition beyond that achievable by conventional mass spectrometers.

Recently, biotechnology firm Medivir AB, Huddinge, Sweden, used the Synapt HDMS system to hasten its drug discovery efforts in the area of protease inhibitor research to eventually help patients who suffer with herpes, hepatitis C, HIV, osteoporosis, osteoarthritis, and high blood pressure. Installed in June 2007, the Synapt HDMS system is yielding significant information that is assisting with the decision-making process and helping Medivir reach its drug discovery milestones.

“Medivir is constantly reviewing opportunities to improve research output both in terms of quantity and quality, says Bertil Samuelsson, VP of discovery research at Medivir. “The Waters Synapt HDMS instrument has been an important vehicle to increase research output and to provide high quality support for the discovery disciplines.”

“The sensitivity of this instrument is significantly greater than the average quadrupole TOF instrument and gives us more complete metabolic profiles,” says Kurt Benkestock, senior research scientist in the analytical chemistry dept., who supports many of Medivir’s research efforts. “At one time, we relied on outside resources to give us the information we needed. Now with the Synapt HDMS system, we can acquire the data we need in-house and move candidates into advanced development phases faster.”

Benkestock’s plans for the Synapt HDMS system are far-ranging, including structure elucidation of metabolites and impurities, ligand screening during early drug discovery, protein and peptide work, protein identification, studying post-translation modifications, and target characterization.

Improving proteomics research

Additionally, Alison Ashcroft and Sheena Radford at the Univ. of Leeds’ Astbury Centre for Structural Molecular Biology, UK, are using the Synapt HDMS system to study the function of biological molecules. They successfully separated and analyzed various folded forms of two important proteins, cytochrome c and beta-2-microglobulin, an achievement they hope will lead to a more complete understanding of the biological processes responsible for amyloid fibril formation, bacterial pilus aggregation, and virus capsid assembly—all associated with debilitating diseases.

Proteins are carefully folded, 3-D long-chain molecules assembled by the human body. When properly folded, they regulate normal bodily functions. Several high profile diseases, including Alzheimer’s, Creuzfeldt-Jakob’s, and Parkinson’s, can develop when certain proteins normally folded into a particular shape become misfolded, causing a chain of events that can lead to self-aggregation and amyloid fibril formation.

Ashcroft and Radford are studying beta-2-microglobulin to try to understand how it forms such fibrils which accumulate in the joints of dialysis patients and which are associated with dialysis-related amyloidosis. A detailed understanding of these processes at the molecular level will aid in the design of therapeutics.

Improving on conventional methods

Waters’ Synapt HDMS system employs technology to analyze samples and differentiate sample ions by their size, shape, and charge, as well as their mass. Image: Waters Corp.

As a tool, conventional MS is an excellent method for differentiating between proteins having different masses. However, as different conformers, or folded forms, of a particular protein have identical masses, it is virtually impossible to differentiate them by conventional means. That’s where the Synapt HDMS system, and its embedded ion mobility technology, has helped Ashcroft.

“A protein can fold into a very compact 3-D structure, or under certain conditions it can unfold into a more expanded structure,” explains Ashcroft.

“Even though these different 3-D structures have the same mass and mass-to-charge (m/z) ratio, the ion mobility capability of the Synapt HDMS system can separate them and tell you how much of the protein is in its folded form vs. the unfolded form. And as the cross-sectional areas of the two protein conformers are different, by being able to separate them by shape, the Synapt HDMS system allows us to distinguish between the various protein forms.”

The Synapt system has provided the lab with new insights into the assembly process. “It’s adding a new dimension to our research,” adds Ashcroft. “We can now quantify the amount of protein that is in its native state and the amount that is unfolded and partially folded. We can also monitor which particular conformers are consumed during the assembly process. This is providing important new insights and detail into how biomolecules work at the molecular level.”

Managing terabytes of scientific data

Given its complement of five different types of mass spectrometers, funded by the Biotechnology and Biological Sciences Research Council (BBSRC) and the Wellcome Trust, managing data is a huge challenge for Ashcroft’s laboratory. To manage data files more efficiently, the laboratory has chosen Waters NuGenesis Scientific Data Management System (SDMS).

“Backing up data daily to DVDs was getting out of hand,” explains Ashcroft. “The SDMS takes data once a day from five mass spectrometers and backs it up automatically, and our post-graduate students and post-docs can look at the data directly from their office computers.”

It’s important to archive data because the government funding bodies request that data is stored for five or 10 years from the date it was created. Graduate students take four years to get their Ph.D.s, so they need the data accessible for four years or more, especially if they are writing papers after getting their Ph.D.

“People that aren’t analytical chemists tend to think of a mass spectrometer as a sophisticated weighing machine,” continues Ashcroft. “Often they don’t realize that you can look at a protein’s functionality and behaviour with the instrument and they can be quite surprised when they find out.”

Insight into infectious diseases

UC Davis uses the Waters LCT Premier XE mass spectrometer in their Waters Corp. Facility for Advanced and Integrated Mass Spectrometry. Image: Waters Corp.

In December 2007, the Univ. of California, Davis (UC Davis) dedicated the new Waters Corp. Facility for Advanced and Integrated Mass Spectrometry within Julie Leary’s laboratory for mass spectrometry. It has been equipped with a Synapt HDMS and LCT Premier XE electrospray TOF mass spectrometer.

Leary, professor of molecular biology and chemistry, together with collaborators at UC Davis, UC Berkeley, and Stanford Univ., all of Calif., is currently using the Synapt HDMS system in her laboratory to study eukaryotic translation, the process by which messenger RNA is translated into proteins within the human 40S ribosome, to better understand how infectious diseases like hepatitis C and polio virus are transferred to humans.

“A key to our current research is to understand the shape of large, complex proteins as they come in contact with each other and with messenger RNA,” says Leary. “Are these proteins folded or are they elongated? What is the topography of the complex?”

For example, Leary’s lab is also using the Synapt to measure the eukaryotic initiation factor, an intact 13 protein complex from human tumor cells which are used to study protein translation.

“We are investigating how the individual protein subunits conform as they come off the intact complex,” explains Leary.

“Biological molecules are dynamic and ever-changing, presenting scientists with a challenge when we try to fully characterize or describe them. We hope that by tackling the issue of molecular shape during eukaryotic translation, we can better understand the process and in turn persuade others to develop effective treatment options.”

—Martha Walz

Resources

• Medivir AB, Huddinge, Sweden, 46 8 5468 3100,
www.medivir.com

• Univ. of California, Davis, 530-752-1011,
www.ucdavis.edu

• Univ. of Leeds Astbury Centre for Structural Molecular Biology, UK, 44 113 243 1751,
www.astbury.leeds.ac.uk

• Waters Corp., Milford, Mass., 800-252-4752,
www.waters.com