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Combinatorial Chemistry &
High Throughput Screening
ISSN: 1386-2073
Combinatorial Chemistry &
High Throughput Screening
Volume 10, Number 3, March 2007
Contents
Reproducibility and Scalability of Solvent-Free
Microwave Assisted Reactions: From Domestic Ovens to Controllable
Parallel Applications Pp. 163-169
Ángel Díaz-Ortiz, Antonio de la Hoz, Jesús
Alcázar, José R. Carrillo, María A. Herrero,
Alberto Fontana and Juan de Mata Muñoz
[Abstract]
Selection of New Chromogenic Substrates of Serine
Proteinases Using Combinatorial Chemistry Methods
Pp. 171-180
Magdalena Wysocka, Bozena Kwiatkowska, Marek Rzadkiewicz,
Adam Lesner and Krzysztof Rolka
[Abstract]
Homogenous Time Resolved Fluorescence Assay to Identify
Modulators of Cap-Dependent Translation Initiation
Pp. 181-188
Regina Cencic, Yifei Yan and Jerry Pelletier
[Abstract]
Target Specific Compound Identification Using a Support
Vector Machine Pp. 189-196
Dariusz Plewczynski, Marcin von Grotthuss, Stephane A.H.
Spieser, Leszek Rychlewski, Lucjan S. Wyrwicz, Krzysztof Ginalski
and Uwe Koch
[Abstract]
Directed Evolution of Oxygenases: Screening Systems,
Success Stories and Challenges Pp. 197-217
Kang L. Tee and Ulrich Schwaneberg
[Abstract]
Fluorous-Enhanced Multicomponent Reactions for Making
Drug Like Library Scaffolds Pp. 219-229
Wei Zhang
[Abstract]
Analysis of Similarity/Dissimilarity of Long DNA Sequences
Based on Three 2DD-Curves Pp. 231-237
Yusen Zhang and Wei Chen
[Abstract]
Abstracts
[Back to top]
Reproducibility and Scalability of Solvent-Free
Microwave Assisted Reactions: From Domestic Ovens to Controllable
Parallel Applications
Ángel Díaz-Ortiz, Antonio de la Hoz, Jesús
Alcázar, José R. Carrillo, María A. Herrero,
Alberto Fontana and Juan de Mata Muñoz
The heating of different parallel arrays in domestic ovens
offers the possibility to perform multiple reactions in one
irradiation experiment, blending the advantages of microwave
heating technology and parallel chemistry. However, they are
usually performed without an appropriate temperature control;
thus, reproducibility becomes a major issue limiting the application
of such reactions. This is exemplified when working at a different
scales or using different instruments. For the first time
a typical solvent-free reaction described in a domestic oven
has been reproduced in monomode reactor, scaled up in a controlled
multimode oven and reproduced in parallel, 24 reactions were
carried out in a well plate. Parallel reactions were performed
in a Weflon®
multiwell plate to assure identical conditions for each individual
reaction. As many reactions under microwave irradiation have
been performed in solvent-free conditions, this result opens
new possibilities in reproducibility, scalability and combinatorial
chemistry and permits to take advantage of many synthetic
procedures described in domestic ovens.
[Back to top]
Selection of New Chromogenic Substrates of Serine
Proteinases Using Combinatorial Chemistry Methods
Magdalena Wysocka, Bozena Kwiatkowska, Marek Rzadkiewicz,
Adam Lesner and Krzysztof Rolka
Chemical synthesis, physicochemical characterization and kinetic
investigations of a tetrapeptide library of chromogenic substrates
containing the amide of 5-amino-2¬nitrobenzoic acid (Anb5,2-NH2)
at their C-termini are reported. Anb5,2-NH2
served as a chromophore released upon enzymatic action. The
library consisting of 9567 peptides was synthesized using
the portioning-mixing method and was screened against bovine
α-chymotrypsin
and human leukocyte elastase in solution applying an iterative
approach. The selected chromogenic substrates were resynthesized
and further modified at their N- and C-termini. Finally, two
sequences, Z-Phe-Ala-Thr-Tyr-Anb5,2-NH2
and Z-Phe-Phe-Pro-Val-Anb5,2-NH2, were
obtained as highly specific substrates for bovine α-chymotrypsin
and human leukocyte elastase, respectively. The method of
synthesis and selection of chromogenic substrates of serine
proteinases described herein is straightforward and can be
applied to design substrates for other proteases.
[Back to top]
Homogenous Time Resolved Fluorescence Assay to Identify
Modulators of Cap-Dependent Translation Initiation
Regina Cencic, Yifei Yan and Jerry Pelletier
Eukaryotic initiation factor (eIF) 4F plays a key role in
recruiting 40S ribosomes and associated factors to mRNA templates
during translation initiation. The function of this heterotrimeric
complex is to deliver an RNA helicase to the 5’ cap
proximal region of mRNAs in preparation for ribosome binding.
To study the interaction between subunits of this complex,
as well as identify small molecules that could interfere with
their association, we developed a time resolved fluorescence
assay that allows monitoring of interactions between two subunits
of eIF4F. We have performed a small molecule chemical screen
of >73,000 compounds using this assay.
[Back to top]
Target Specific Compound Identification Using a Support
Vector Machine
Dariusz Plewczynski, Marcin von Grotthuss, Stephane A.H.
Spieser, Leszek Rychlewski, Lucjan S. Wyrwicz, Krzysztof Ginalski
and Uwe Koch
In many cases at the beginning of an HTS-campaign, some information
about active molecules is already available. Often known active
compounds (such as substrate analogues, natural products,
inhibitors of a related protein or ligands published by a
pharmaceutical company) are identified in low-throughput validation
studies of the biochemical target. In this study we evaluate
the effectiveness of a support vector machine applied for
those compounds and used to classify a collection with unknown
activity. This approach was aimed at reducing the number of
compounds to be tested against the given target. Our method
predicts the biological activity of chemical compounds based
on only the atom pairs (AP) two dimensional topological descriptors.
The supervised support vector machine (SVM) method herein
is trained on compounds from the MDL drug data report (MDDR)
known to be active for specific protein target. For detailed
analysis, five different biological targets were selected
including cyclooxygenase-2, dihydrofolate reductase, thrombin,
HIV-reverse transcriptase and antagonists of the estrogen
receptor. The accuracy of compound identification was estimated
using the recall and precision values. The sensitivities for
all protein targets exceeded 80% and the classification performance
reached 100% for selected targets. In another application
of the method, we addressed the absence of an initial set
of active compounds for a selected protein target at the beginning
of an HTS-campaign. In such a case, virtual high-throughput
screening (vHTS) is usually applied by using a flexible docking
procedure. However, the vHTS experiment typically contains
a large percentage of false positives that should be verified
by costly and time-consuming experimental follow-up assays.
The subsequent use of our machine learning method was found
to improve the speed (since the docking procedure was not
required for all compounds from the database) and also the
accuracy of the HTS hit lists (the enrichment factor).
[Back to top]
Directed Evolution of Oxygenases: Screening Systems,
Success Stories and Challenges
Kang L. Tee and Ulrich Schwaneberg
The field of directed evolution of oxygenases (mono-, di-
and epoxygenases) is rapidly advancing as an increasing number
of success stories indicate. A significant number of screening
systems have been developed to specifically improve oxygenase
properties. Oxygenases will become very valuable biocatalysts
for synthetic applications in industry when stability, cofactor
and activity properties match industrial demands. This review
summarizes screening systems and principles of screening systems
that have been used for directed evolution of oxygenases.
Sections on mutagenic conditions, mutant library size and
property improvements provide a comprehensive picture on performance
and limitations of current directed evolution methodologies
for oxygenases. A discussion of challenges in the directed
evolution of oxygenases for industrial exploitation concludes
this review.
[Back to top]
Fluorous-Enhanced Multicomponent Reactions for Making
Drug Like Library Scaffolds
Wei Zhang
Multicomponent reactions (MCRs) generate multiple bonds in
a single reaction process, which is highly efficient to construct
relatively complex molecules. Conducting post-MCR modification
reactions further increases the molecular complexity and diversity.
MCR has become a powerful approach to make drug-like molecules
in lead generation chemistry. In fluorous MCR (F-MCR), one
of the starting materials is attached to a fluorous tag and
used as the limiting agent. After the MCR, the fluorous component
is fished out from the reaction mixture and used for post-MCR
modifications. The fluorous tag can be finally removed in
traceless fashion by displacement or cyclization reactions.
Unique fluorous technology such as fluorous solid-phase extraction
(F-SPE) facilitates the separation process. Other techniques
such as microwave irradiation and plate-to-plate SPE can also
be used to make the F-MCR even more efficient. Syntheses of
unique heterocyclic and natural product-like library scaffolds
using Ugi/de-Boc/cyclization, MCR/Suzuki coupling, and [3+2]
cycloaddition/de-tag/cyclization protocols are described in
this paper.
[Back to top]
Analysis of Similarity/Dissimilarity of Long DNA Sequences
Based on Three 2DD-Curves
Yusen Zhang and Wei Chen
Some 2D graphical representations of DNA sequences have been
reported by several authors, which give visual characterizations
of DNA sequences. In this paper, we present a new 2D graphical
representation of DNA sequences without degeneracy. Furthermore,
we propose two methods for the visualization and analysis
of long DNA sequences.
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