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History of the YCC/Keck Mass Spectrometry Resource
History of the YCC/Keck Mass
Spectrometry Resource
The history of mass spectrometry in
the Yale Medical School has its origin in 1965 in the laboratory of Dr.
Sandy Lipsky in the Department of Medicine. Professor Lipsky was an
expert in gas chromatography which is a technique that he used to
separate and analyze complex mixtures of lipids in the gas phase and
which relies on relatively high temperatures to volatilize the lipids.
To facilitate analysis and identification of the lipids, he coupled a
mass spectrometer detector to a gas chromatograph – becoming one of the
early laboratories to utilize the new technique of gas
chromatography-mass spectrometry (GC-MS). At that time this was not an
easy technique to implement since it required interfacing a high
pressure separation technique (GC) to a high vacuum and high voltage
instrument (MS). NASA provided ~$125,000 to purchase an AEI MS-9 mass
spectrometer and Dr.'s Sandy Lipsky and Walter McMurray used this
instrumentation to analyze Apollo 11 lunar samples. As part of this
research effort they developed the technique to electrically record
rapidly scanned mass spectra. Prior to this advance, spectra were
recorded on photographic paper or photographic plates and the masses
were determined by manually counting the peaks in the spectrum. The
computer was an IBM 1800 which occupied about 15 ft of wall space.
Simultaneously with the development of the GC-MS technique, Professor
Lipsky and Professor Csaba Horvath (the Roberto C Goizueta Professor)
in the Department of Chemical Engineering, Yale University were
developing the technique of High Pressure Liquid Chromatography (HPLC)
which is now one of the most widely used techniques for separating and
analyzing complex mixtures of proteins, peptides, amino acids, and
other biomolecules.
The Yale Comprehensive Cancer Center Mass Spectrometry Resource was
founded in January, 1984 by Professor Alan Sartorelli (who was then
Director of the YCC) with Dr. Walter McMurray as the Resource Director.
At this time the Mass Spectrometry Resource had only one instrument
which was a VG 16F single focusing mass spectrometer equipped with a
Varian gas chromatograph. The data system for this instrument was a
Digital PDP-8 computer. A grant submitted to the NIH under their
DRR-BRS Shared Instrumentation Program was funded for $300,000 (the
maximum amount permitted) in December, 1984. These funds, together with
approximately $225,000 provided by the YCC, was used to purchase a VG
ZAB-SE. This double focusing mass spectrometer was equipped with gas
and liquid chromatographic interfaces and used a Digital PDP11-24
computer as a data station. Due to extensive renovation of the
laboratory space in which the ZAB was located, installation was not
completed until March, 1987. The laboratory space was actually located
at 385 Congress Avenue which is now the site of the new Congress Avenue
Building. The funding for these renovations was provided by Yale
University. During the next few years the ZAB was upgraded twice
through funds acquired from NIH Small Instrument Grants.
In the early 90's the ZAB was moved (which was not an easy task as the
magnet weighed a ton) from 385 Congress Ave to newly renovated space in
the basement of Winchester, two floors below the Cancer Center’s
offices. The funds for these renovations were also provided by the
University. The move to the new space was accompanied by a second
NIH-BRS Shared Instrumentation Award for $365,000 for the purchase of a
VG Quattro triple quadrupole mass spectrometer equipped with an
electrospray ion source. Electrospray, developed by Professor. John
Fenn and graduate student Craig Whitehouse, Department of Chemical
Engineering, Yale University, revolutionized (concurrently with the
development elsewhere of Matrix Assisted Laser Desorption Ionization (MALDI))
the mass spectral analysis of biological molecules - particularly
proteins. The data system for this instrument was a PC using Windows.
In 1993 the Keck Laboratory Mass Spectrometry Resource was founded by
Kathryn Stone and was equipped initially with a loaner TofSpec MALDI-MS
from Micromass, Inc which was replaced in 1995 by a research-grade
TofSpec SE instrument funded with a joint NIH/NSF Shared
Instrumentation Grant (RR09167/BIR9319358). The following year the Keck
Laboratory MS Resource was awarded a grant from the Howard Hughes
Medical Institute to fund the purchase of an LCQ, an ion trap mass
spectrometer equipped with an electrospray ion source and interfaced to
a liquid chromatograph.
From the late 1980's there had always been close interactions between
the YCC Mass Spectrometry Resource and several of the Resources that
make up the Keck Laboratory. One nice example of this collaborative
effort were the studies these two laboratories carried out to solve the
structure of the "unknown" Standard Test Peptide 3 (STP-3) that was
specifically designed (e.g., it included a branched structure in which
the two branches were of unequal lengths, an acylated epsilon lysine
amino group, a disulfide linkage, two methionine residues susceptible
to air oxidation, and whose sequence was chosen to be refractory to
enzymatic cleavage) to challenge state-of-the-art capabilities of
protein chemistry and mass spectrometry laboratories around the world.
This sample was distributed to 180 registrants who requested it in
conjunction with the 1988 Protein Society Meeting Together, the YCC
Mass Spectrometry and Keck Laboratories produced one of only three
correct answers for the structure of this challenge peptide - with
these studies being described in a publication by Elliott et al ( 1 ).
The YCC and Keck Laboratory Mass Spectrometry Resources formally merged
in July, 1998 - with the Directors of each Resource (Walter McMurray,
Ph.D. and Kathryn Stone B.S. respectively) becoming Co-Directors of the
joint Resource. The first instrumentation grant application submitted
by the joint Resource resulted in a $630,000 award (NSF987111) that was
made jointly by the National Science Foundation and the Howard Hughes
Medical Institute. This award funded the purchase of one of the first
quadrupole/time-of-flight mass spectrometers (a Micromass Q-Tof) to be
placed in academia. This instrument has an electrospray source and is
still in continual use for determining molecular weights of proteins
and other biomolecules (which often exceed 100 kDa), the exact mass of
small (<1000 Da) synthetic intermediates in chemical syntheses,
and to determine amino acid sequence tags for peptides obtained
(usually) from gel-separated proteins submitted for protein
identification.
The NIH Shared Instrumentation Grant application (RR015837) submitted
in 2001 by the joint Resource was ranked highest in the country and
provided $497,200 in funding that was used towards the purchase of two
mass spectrometers. The first system purchased was a Micromass MALDI-R
mass spectrometer system that forms the basis for high throughput
protein identification and peptide disease marker discovery services
that are now available This highly automated system includes a
Micromass MassPrep robot which automatically manipulates gel slices in
96-well plates from tryptic digestion through extraction to spotting
onto the MALDI target plate. Under computer control, spectra are
acquired, processed, and database searched to identify the protein(s)
in the gel. Recently, HHMI awarded the Resource $140,000 to upgrade
this system to the MALDI-L/R configuration which will substantially
extend the molecular weight range past the current limit of about
3,500. The second system that was partially funded by this NIH Shared
Instrumentation Grant was a very high throughput Sequenom SNP
genotyping system that utilizes a single base extension/MALDI-MS
detection methodology and is capable of analyzing >20,000
genotypes/day.
.
The most recent grant application submitted by the YCC/Keck MS Resource
was in response to a one-time RFA for the extremely competitive NIH
High End Instrumentation Program. The resulting $1.4 million dollar
award is the largest instrumentation grant received by the Yale School
of Medicine and it will fund the purchase of an extremely advanced,
Fourier Transform Ion Cyclotron Resonance Mass Spectrometer (FTICR-MS)
which has uniquely high mass resolution and accuracy. This instrument,
which will be equipped with both MALDI and electrospray sources, is
expected to be used for determining (with extremely high mass accuracy)
masses on a wide range of proteins and other biomolecules and for
analyzing extremely complex digests of whole cell protein extracts for
the purpose of identifying protein post-translational modifications.
We believe the merging of the YCC and Keck Laboratory Mass Spectrometry
Resources, which are described in more detail at
http://keck.med.yale.edu/prochem/, has been an unqualified
success that has resulted in substantially strengthening the mass
spectrometric support that is available to members of the Yale Cancer
Center and to the Yale Community. That the level of support provided by
this Resource is extremely high is suggested by its widespread use by
non-Yale investigators and also, by comments made by NIH and NSF Study
Panels on its grant applications. Hence, in fiscal year 2001 the YCC/Keck
MS Resource provided analyses for 65 Yale and 234 non-Yale principal
investigators with the latter at 110 institutions around the world
including Argonne Natl. Lab, Brookhaven Labs, Dartmouth, Duke, Harvard,
Johns Hopkins, MIT, NIH, Rice, Stanford, and Washington U. The
following are comments made by NIH Study Panels on the two most recent
awards made to the YCC/Keck MS Resource:
NIH, RR15837, "High Throughput MALDI Mass Spectrometer System",
(2001), PI: K. Williams: “This is an outstanding proposal from a
premiere core laboratory at Yale.” "The facility has a long track
record of superior service and efficient fiscal management." "The Keck
Biotechnology Resource Laboratory is arguably one of the best
facilities of its kind in the nation in terms of support and expertise.
The large number of researchers who annually make use of this facility,
both from the Yale campus and nationwide, is testament to the
significance of the facility for crucial support of biomedical and
biotechnology research." "Dr. Williams is the founder and current
Director of the Keck Lab, which he has organized and built into one of
the best Biomedical Resource Laboratories in the country. Dr. McMurray
has extensive experience with mass spectrometers. Kathy Stone has
published many methodological papers (often with Dr. Williams) that
document her commitment to developing practical approaches to the
difficult sample handling protocols that are fundamental to a Core
Laboratory." "The expertise of the laboratory is also evident in the
leading role Dr. Williams and Ms. Stone have played in the development
and growth of the Association of Biomedical Resource Facilities, an
organization that has improved the quality and accessibility of Core
laboratories around the country -- often emulating the Keck model at
Yale."
NIH, RR017266, "FTICR Mass
Spectrometer", (2002), P.I. K. Williams. "Strengths
brought up during the panel discussion included 1) the clear
justification of need - all of the research projects will demand the
capabilities of high field FTICR-MS, 2) the highly meritorious nature
of the biomedical research which will be advanced by the proposed
instrument, 3) the sound institutional commitment and administrative
plan, 4) the key development of up-front separation methodology for
ICAT- related research, and especially, the development of
bioinformatic tools for data interpretation and correlation with
microarray data that is critical for the research projects, and 5) the
impressive track record of the PI's facility, which has a history of
providing first-rate services to support well-funded and interesting
science. Dr. McMurray, who oversees the MS operation, provides
excellent expertise in mass spectrometry. The facility also has
strengths in many other areas, including bioinformatics and separation
science, which should interact well with the new instrument. The
research projects of the major and minor users listed as well as others
not listed is extensive, impressive and well funded. The two main
driving forces of the proposal are relative protein quantification and
post-translational modification determinations. Furthermore, the
development of up-front separation methods for ICAT-like experiments as
well as data-analysis, data management and data-integration strategies,
also well funded, is a real strength of this proposal."
1. Elliott, J., Stone, K., Roberts,
W., LoPresti, M., Crawford, M., Kapouch, J., Jacobsen, E., Williams,
K.R., McMurray, W., Meng, C., Mann, M. and Fenn, J.(1989) Structure of
Symposium Test Peptide-3. In: Techniques in Protein Chemistry. (Hugli,
T.E., ed.) Academic Press, New York, 569-579.
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