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Dear All,
I wonder whether plasma proteins can bind drugs at stoichiometry
greater than 1:1. If so, how high the stoichiometry can be? Any
literature evidence is much appreciated.
Thanks,
Kasiram.
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The following message was posted to: PharmPK
Kasiram,
Yes, any hypothetical drug can bind any hypothetical plasma protein
at higher than unit ratios, but it is always dependent on the surface
and active sites any given protein presents for binding, whether the
protein remains in its native conformation or whether its
conformation has been perturbed (i.e., denatured, cleaved, etc.) AND
it depends on the ionic and/or other associative means any given drug
has available to the proteins in solution.
I can't provide literature references, but binding is often what
drugs and proteins do with each other. There have to be numerous
references available.
Certainly, a chief plague to any bioanalyst begins with the chemical
stability of the drug, then moves on to whether it associates with
proteins or other "surfaces" during sample collection. The success
and particularly the precision of any bioanalysis is directly related
to the success of sample separation and analysis techniques to
measure ALL drug present in a sample, not just the amount available
through inadequate separation.
Best wishes,
Ian Davis
Director, Operations
Strategic Consulting Services
Pharsight Corporation
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Dear Kasiram,
with the available structural data for albumin you can actually see
in the crystal structures that several drugs/molecules can be bound
per albumin molecule.
for fatty acids 7 sites have been clearly identified on albumin:
Bhattacharya AA, Grune T, Curry S (2000) Crystallographic analysis
reveals common modes of binding of medium and long-chain fatty acids
to human serum albumin. J.Mol.Biol.; 303 (5):721-32.
Petitpas I, Grune T, Bhattacharya AA, et al (2001) Crystal structures
of human serum albumin complexed with monounsaturated and
polyunsaturated fatty acids. J.Mol.Biol.; 314 (5):955-60.
But also for drugs up to 3 molecules per albumin have been found:
Ghuman J, Zunszain PA, Petitpas I, et al (2005) Structural basis of
the drug-binding specificity of human serum albumin. J.Mol.Biol.; 353
(1):38-52.
In addition drugs can/will also bind to other plasma proteins, blood
cells etc. due to the high concentrations of many of the binding
sites in plasma/blood, relatively low affinities are needed for a
significant binding.
With regards,
Markus
Markus Weiss, PhD
ED/DMPK/ADME/ADE
Novartis Pharma AG, Werk Klybeck
WKL-135.4.83
CH-4002 Basel
Switzerland
Phone: +41 61 69 61075
Fax: +41 61 69 65146
Email : markus.weiss.at.novartis.com
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The following message was posted to: PharmPK
Thanks Markus, for the information. But, I am more interested in binding
of drugs to plasma proteins, especially binding of basic drugs to
alpha-1-acid glycoprotein. I have seen references suggesting
stoichiometry as high as 1:20. Is it true to have such a high
stoichiometry and, if so, how high can this be?
There are two issues about binding sites on protein - different classes
of sites and number of sites per class. I am interested in both and
wonder whether there is any means other than conventional scatchard plot
analysis to know the number of binding sites. For eg. is it possible to
know from the crystal structure?
I appreciate any inputs from the group.
Kasiram
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Dear Kasiram,
for AGP there is no experimental structure, it is highly glycosylated
and can therefore not be easily crystallized. There is however a
model based on sequence similarity, AGP is an outlier lipocalin and
should form a beta-barrel (see Kopecky et al 2003 Biochem Biophy
Research Communications). 20 binding sites seems rather high to me,
but I guess you can contact the authors of the above paper and they
can tell you how big the inside of the barrel is, and how many
compounds they think would fit or how many binding sites they expect.
I remember from this or another publication (Flower et al 2000
Biochim Biophys Acta) that people were talking about three sites?
I guess Biacore could be an elegant system to look at stoichimetry.
Regards,
Markus
Markus Weiss, PhD
ED/DMPK/ADME/ADE
Novartis Pharma AG, Werk Klybeck
WKL-135.4.83
CH-4002 Basel
Switzerland
Phone: +41 61 69 61075
Fax: +41 61 69 65146
Email : markus.weiss.at.novartis.com
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The following message was posted to: PharmPK
Dear Kasiram
I agree with Markus, a stochiometry higher than 1 for albumin is
frequent
(Bree et al Biochem Pharmacol 38; 753-58, 1989), it is the same for the
lipoproteines for which the number of binding sites can be higher
than 10
one tends then rather towards a nonsaturable binding on the other
hand for
the orosomucoide (AAG) the stochiometry is always 1/1
Francoise
Francoise BREE
Test Facility Management
BIOPREDIC
Parc d'affaire de la Breteche
Batiment B1
35760 Saint Gregoire
France
francoise.bree.-at-.biopredic.com
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