Molecular Recognition Laboratorium

Contact Information

Molecular Recognition Laboratorium,
Institute für medizinische Immunologie
CHARITÉ - UNIVERSITÄTSMEDIZIN BERLIN
[show][hide]

	Hessische Str. 3-4
	D-10115 Berlin, Germany
	
	phone  +49-30-450 524092 
	fax    +49-30-450 524942 
	mail   annette.hayungs@charite.de
 	web    charite.de

last change on 01.04.2010

Group Leader

Rudolf Volkmer [show][hide]

	Tel.:	+49 30 450 524 267
	Fax:	+49 30 450 524 942
	E-Mail:	rve(at)charite.de
	
	

Group Members

• Magdalena Czuban, Dipl. Biol. [show][hide]

	Tel.:	+49 30 450 xxx xxx
	Fax:	+49 30 450 524 942
	E-Mail:	x(at)charite.de
	 	

• Mercedes Gonzáles, Studentin [show][hide]

	Tel.:	+49 30 450 xxx xxx
	Fax:	+49 30 450 524 942
	E-Mail:	x(at)charite.de
	 	

• Anja Heiduk, Dipl. Biol. [show][hide]

	Tel.:	+49 30 450 xxx xxx
	Fax:	+49 30 450 524 942
	E-Mail:	x(at)charite.de
	 	

• Simone Jagdhuber, Studentin [show][hide]

	Tel.:	+49 30 450 xxx xxx
	Fax:	+49 30 450 524 942
	E-Mail:	x(at)charite.de
	 	

• Ines Kretzschmar, TA [show][hide]

	Tel.:	+49 30 x
	Fax:	+49 30 x
	E-Mail:	x(at)charite.de
	 	

• Christiane Landgraf, TA [show][hide]

	Tel.:	+49 30 450 524 xxx
	Fax:	+49 30 450 524 942
	E-Mail:	chl(at)charite.de
	 	

• Eric Moinet, Student [show][hide]

	Tel.:	+49 30 450 524 xxx
	Fax:	+49 30 450 524 942
	E-Mail:	x(at)charite.de
	 	

• Víctor Tapia - Doktorand [show][hide]

 	Tel.:	+49 30 450 524 285
	Fax:	+49 30 450 524 942
	E-Mail:	victor.tapia(at)charite.de; ve.tapia.m(at)gmail.com
	 	

Research interest

The AG Volkmer emerges from a strong training in peptide chemistry and cultivates its expertise in the synthesis and preparation of peptide/peptoide probes, either as core service or to meet the demands of our own research in cellular biology and molecular medicine. The objectives of our research principally spread around three main topics: profiling the binding specificity of protein recognition modules, analyzing the diagnostic potential peptide-based analyte capture assays, and exploring the potential of peptide/peptoide probes to define or modulate specific therapeutic strategies.

Profiling the binding specificity of protein recognition modules

Protein recognition modules (PRM) are non-catalytic domains of protein structure dedicated to read molecular motifs of primary structure and post-translational modifications of proteins. Reading or recognition is not restricted to protein motifs, as shown by the emerging field of epigenetics. It is clear that methylation and other signatures on DNA are also recognized by modular structures of nuclear factors. The tinkering of evolution has repeatedly duplicated and diverged different structural modules resulting in several homology families with some degree of functional conservation. This is observed as regular expressions of short linear motifs that can be recognized by a PRM family and by specific affinity traits of individual PRMs. We aim to identify key events in cellular processes of information reading and transduction. Success in such aim translates to engineering congruent interaction networks, complementing drug efficiency and designing new therapeutic strategies, as has been shown by our group with several domains, i.e. WW domains in X-linked intellectual disorders and mechanosensing, SH3 domains in endocytosis, as well as PDZ domains in cystic fibrosis.

Analyzing the diagnostic potential of peptide-based analyte capture assays

We intend to focus away from single biomarker for diagnosis and use either validated collections or agnostic collections of peptide probes to analyze complex biological samples. The former approach relies on the use of peptide probes known to interact with one or multiple target biomarkers for a specific pathology, the later one relies on extensive stochastic peptide probes to capture eventually unknown analytes in the sample that can be used as a diagnostic pattern of detection signals upon a multiplex binding assay. The development of these approaches can be estimated from reports of our group and cooperation partners inside as well as outside RCIS. These reports show the difficulties and potential of the use of peptide probes to analyze blood samples and diagnose hypersensitivity, autoimmune responses, and immunological responses to infection.

Exploring the therapeutic potential of peptide/peptoid probes

The flag ship of this research venue derives from the unique expertise for generating immobilized but C-terminal exposed peptide probes and accurate experience with PDZ specificity profiles. A peptide probe has been design to selectively inhibit the CFTR–CAL interaction —relevant in cystic fibrosis— without affecting the biologically relevant PDZ competitors NHERF1 and NHERF2. In this venue of research we also explore the potential of cell penetrating peptides to be applied as vector to transport drug probes, such as the CFTR-CAL inhibitor, across the cell membrane.

Technological Development of the Peptide Array Technologies

by Victor Tapia

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The combination of SPOT peptide synthesis (figure A, steps 1 to 4) with appropriate immobilization techniques on glass supports (figure A, steps 5 and 6) is wide spread. The SPOT technology provides low-scale but high-throughput synthesis, while immobilization of pre-synthesized peptides offers the benefit of a "chemical" purification step and flexible array design. Additionally, the glass support is compatible with fluorescence detection and offers the possibility to miniaturize binding assays. Beyond economy, the later point is essential for quantitative measurements at the steady-state of binding activity, as has been described [Ekins 1998] and can be proven by the mass-action law.

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The basic point of this technology is the simultaneous display of a
systematic collection of peptides on a planar support, on which numerous
bimolecular interaction assays can be carried out under homogeneous
conditions.

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PEPTIDE ARRAYS IN THE ADVANCEMENT OF PEPTIDE SYNTHESIS

• The development of solid-phase peptide synthesis (SPPS) by Bruce Merrifield [Gutte and Merrifield, 1969; Merrifield, 1965] and adaptions of this procedure [Fields and Noble, 1990] set the chemical ground for innovative technologies to follow.
  
• The development of the “Pin” method by H. Geysen [Geysen, et al., 1984] introduces the array format to peptide synthesis.
  
• Definitive establishment of peptide arrays came along with the development of the SPOT synthesis by Roland Frank [Frank, 1992; Frank, 2002] which simplified chemical synthesis of peptide arrays to the addressable deposition of reagents on a cellulose sheet.
  

Modern peptide synthesis approaches and
molecular biology make peptides accessible in a high degree of
structural diversity. The two greatest drawbacks of synthetic peptide
arrays are peptide length, with a quality threshold between 30 and 50
amino-acids, as well as the restriction to linear motives, since the
mimicry of nonlinear motives with linear peptide constructs is still
under development [Goede, et al., 2005].

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PEPTIDE ARRAYS IN THE ADVANCEMENT OF BINDING ASSAY SYSTEMS

Since the 90s a major aspect of development to achieve the required
sensitivities to analyse biological samples has been the miniaturization
of analytical devices [Ekins, 1998]. It is important to note that
miniaturization is not only a matter of high-throughput and economy.
Miniaturization is an essential factor that should provide saturation of
binding sites under low analyte concentrations without significantly
altering its bulk (or ambient) concentration upon capturing [Ekins, et
al., 1990; Ekins, 1989; Joos, et al., 2002; Templin, et al., 2002].

• In this sense, the first application of a peptide microarray device in 1991, anticipating even the application of cDNA arrays, achieved already the impressive feature density of about 1024 peptides in 1.6 cm2 by means of in situ light-directed parallel synthesis [Fodor, et al., 1991].

Several methods available to generate peptide arrays on planar solid surfaces offer a range between...

• 16 peptides per cm2, in the case of SPOT macroarrays [Reimer, et al., 2002; Schutkowski, et al., 2004],
• to 2000-4000 peptides in 1.5 cm2, in the case of microarrays generated by digital photolithography [El Khoury, et al., 2007; Gao, et al., 2004; Pellois, et al., 2000; Pellois, et al., 2002].

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SOURCES: For references to citations above see

• Tapia, V.E., Ay, B., Volkmer, R., 2009. Exploring and Profiling Protein Function with Peptide Arrays, in: Marina Cretich, Chiari, M. (Eds.), Peptide Microarrays. Humana Press, Totowa, NJ, pp. 3–17.
• Tapia, VE & R Volkmer, 2009. Steady State Analysis of Peptide Array-based Binding Assays [1]

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