Although I studied physics and mathematics until the master's level, I always knew
that I wanted to get into biology, whose golden age I was sure was happening in my
lifetime. When the opportunity came knocking – in the shape of John Fenn, who was
spending a sabbatical at the University of Göttingen – I did not hesitate to take
up his offer to come with him to Yale for my PhD. John was working on a far-out idea:
that one could ionize biomolecules by applying an electric field to the outlet of
a needle through which a liquid was flowing. At first there was little interest in
‘electrospray’, but as soon as it became clear that large, intact proteins survive
this process (Mann et al, 1989; Meng et al, 1988), the community was hooked. This
included the Nobel Prize committee, who gave John a share of their prize for chemistry
in 2002.
»I took one of the most ambitious and stressful decisions of my professional life:
I bet everything on the success of mass spectrometry-based protein characterization.«
Electrospray has been my mainstay ever since. The only interruption was as a post-doc,
which I did in Denmark – following my Danish wife, whom I had met at Yale. Peter Roepstorff's
group taught me a lot about protein chemistry but they had not acquired electrospray
yet, so I missed the initial wave of discoveries done with our new technology! This
turned out to be a blessing in disguise, however, because it motivated me to develop
software instead. Specifically I attacked the problem of how to identify peptides
in sequence databases given a minimal amount of mass spectrometric information. When
in 1992 I was offered a group leader position at the European Molecular Biology Laboratory
(EMBL), I took one of the most ambitious and stressful decisions of my professional
life: I bet everything on the success of mass spectrometry (MS)-based protein characterization.
At the time, despite much talk, MS had not become a serious tool for high sensitivity
protein detection in biology, which was dominated by chemical methods such as Edman
degradation instead. Fortunately, we soon made two crucial breakthroughs: Matthias
Wilm devised nanoelectrospray, a miniaturized and highly sensitive form of electrospray,
and Andrej Shevchenko developed protocols for liberating proteins from gels so that
they could be analysed by MS. Together with our novel bioinformatic algorithms (Mann
& Wilm, 1994), and the beginning genome sequencing efforts, this enabled identification
of minute amounts of protein – far exceeding the capabilities of Edman sequencing
and firmly establishing mass spectrometry in molecular biology research (Wilm et al,
1996). Among the prize catches of these years was the identification of caspase-8
(with Peter Krammer and Vishua Dixit) and the catalytic subunit of telomerase (with
Tom Check). I am sure that these successes would not have happened without the high
pressure environment at EMBL. Together with complementary work by the Yates and the
Aebersold groups, these were the first steps of mass spectrometry in biological research,
which eventually led to its establishment as the standard tool for protein identification.
After EMBL, my move back to Denmark (and reunion with my family) was made possible
by the Danish Natural Research Council. The next 7 years were very fruitful and our
group developed many of the technologies that are now in general use – including the
method of stable isotope labelling by amino acids in cell culture (SILAC; Ong et al,
2002), which has become a gold standard in quantitative accuracy for proteomics. I
continued to be involved in programming too, which gave us quite a competitive edge
in devising new strategies to retrieve biologically meaningful data from proteomics.
During these years we started to apply proteomics to define the members of protein
complexes. First examples included the U1 subunit of the yeast spliceosome (with Reinhard
Luehrmann), the entire human spliceosome (with Angus Lamond) and the centrosome (with
Erich Nigg). Interaction and organellar proteomics continue to be some of the most
promising areas for proteomics and together with Tony Hyman in Dresden we are currently
pursuing a large-scale effort to produce a high quality human interactome. We also
used proteomics to study mechanisms of growth factor stimulation of SILAC labelled
cells. By precipitating and then quantifying tyrosine-phosphorylated proteins at different
time points, we could follow the signal as it spread to many and diverse substrates
(Blagoev et al, 2004). Some of the proteins that we cloned at the time have become
quite important in the growth factor signalling field. Several years later, we extended
these experiments to generate a first global picture of ‘early information processing’
by the cell in response to growth stimulus (Olsen et al, 2006).
In 2005, I moved to the Max-Planck Institute of biochemistry in Martinsried (Munich)
where we diversified into a variety of biological and medical directions while keeping
our core emphasis on instrumentation and bioinformatics. The Max-Planck institutes
are fantastic environments for science in general and for our work in particular.
Here, my group is able to work on the entire chain of proteomics – from technologies
of proteomic sample preparation, through chromatographic and mass spectrometric innovations
to methods in computational proteomics (Fig 1). Bioinformatics development connected
to the analysis of proteomics data sets has always been a major focus in my group
but in Martinsried it was kicked into high gear by Jürgen Cox. He developed the MaxQuant
suite of proteomics tools that are now widely used by the community to analysed proteomics
datasets (Cox & Mann, 2008). We then apply all these technologies to a wide range
of biological problems; partly to show that proteomics can be a powerful tool in these
different fields.
»The most ambitious project we undertook at our new location was to crack a complete
proteome.«
Figure 1
Workflow of high resolution and quantitative proteomics.
The most ambitious project we undertook at our new location was to crack a complete
proteome. One of the limitations of proteomics had always been that usually only a
small number of proteins were actually identified and quantified. Pre-MS techniques
such as 2D gel electrophoresis usually identified a few dozens of proteins, a far
cry from the thousands of probes that the microarray community was putting on their
chips. To show that the full proteome was amenable to MS analysis, we collaborated
with Tobias Walther (now at Yale), to quantify haploid versus diploid yeast (de Godoy
et al, 2008). Very recently, we have shrunk the analysis time needed to cover nearly
the entire yeast proteome to just a few hours (Nagaraj et al, 2011a). The human proteome
is not quite complete yet, but we are getting very close (Nagaraj et al, 2011b). However,
these studies are just first steps; in the future the task for proteomics will include
detection and quantification of protein isoforms, a nearly complete set of protein
modifications and doing all this as a function of time and cellular localization –
a full program for at least one more generation of proteomics researchers!
»Using a variant of the SILAC technology called ‘spike-in’ or ‘super’ SILAC it is
now possible to measure the proteome of tumour biopsies at great depth and with extremely
high precision.«
Recently, proteomics technologies have become sufficiently evolved that it is now
realistic to measure clinical material. Using a variant of the SILAC technology called
‘spike-in’ or ‘super’ SILAC it is now possible to measure the proteome of tumour biopsies
at great depth and with extremely high precision (Geiger et al, 2010). This is one
of the areas that my group will focus on in the coming years and we already have proof
of principle in defining protein expression signatures in difficult to diagnose lymphoma
subtypes. It is clear that there is now an opportunity to make a great clinical impact
using high resolution and robust proteomics technologies (Fig 2).
Figure 2
Proteomic analysis of breast cancer samples (Tamar Geiger, Jacek Wisniewski and Matthias
Mann).
In conclusion, whenever one really wants to understand biological function, one has
to deal with proteins and mass spectrometry is the method of choice to do this. Clearly,
the fun is just beginning as we now get to apply the tools developed by the community
over the last decades. So, what does the future hold for proteomics and where will
it make unique contributions? Areas to watch include the analysis of complete mammalian
proteomes, including absolute quantification of cellular proteins, increasingly sophisticated
studies of the function of thousands of post-translational modifications as well as
protein interaction studies. Proteomics is starting to be used to probe the effects
of genome variation between humans at the functional level and I predict that this
will be an expanding area. In a slightly longer perspective, proteomics will become
an important basis on which systems biological modelling of the cell will be built
(Cox & Mann, 2011).