beta-lactamase
Bacterial
infections are arguably some of the most serious threats to humankind
to date. The options for treating infections have dwindled
substantially and the reports of antibiotic resistant pathogens are
increasing at an alarming rate. Once potent antibiotics such as
penicillin nowadays are hardly effective by themselves including
against certain strains of Bacillus anthracis, the causative agent of
anthrax. This antibiotic resistance is due in large part to bacterial
beta-lactamases
 
capable of
degrading penicillin-like drugs. A powerful avenue for treating
infections was the administration of a beta-lactamase inhibiting
co-drug in addition to prescribing penicillin-like antibiotics. There
are currently three beta-lactamase inhibitors on the market,
tazobactam, sulbactam, and clavulanic acid each with an annual sales of
close to or over a billion dollar. The bacterial response to this
co-drug combination was not too surprising and beta-lactamase variants
were soon found to confer Detailed structural knowledge on how these
inhibitors function and how mutations in beta-lactamases confer
resistance to these inhibitors is desperately needed. Protein
crystallography has been a tremendous tool to study beta-lactamases
with and without substrates or inhibitors yet the complexity of the
enzymatic degradation pathway of such compounds has often precluded
obtaining clear crystallographic snapshots of reaction intermediates. A
novel solution to this underlying problem of not knowing what
intermediates are formed at what rate in the crystal once soaking is
commenced has been developed by our collaborative team including Drs.
Paul Carey, Marion Helfand, and Robert Bonomo. This innovative
technique, termed Raman crystallography, has led to the identification
of the trans-enamine intermediate peaking at 20-30 minutes inside the
deacylation deficient E166A mutant of SHV-1 beta-lactamase. Our lab
subsequently determined the 1.63 Ang crystal structure of this
intermediate complex for tazobactam yielding a wealth of detailed
information of how this drug inhibits this enzyme resulting in ideas on
how to rationally improve this drug (tazobactam has an
annual sales of close to a billion dollars in the US).
Click here to view structure or download coordinates.
This work was
published in Biochemistry
Based on this
tazobactam complex, we have designed a novel inhibitor SA2-13 with an
intend to stabilize the trans-enamine intermediate. 
Our designed SA2-13 compound yielded a
10-fold improvement of the longevity of the trans-enamine intermediate
and, unlike the starting tazobactam compound, SA2-13 could now readily
be trapped in wt SHV-1 crystals.
This work was recently published in JACS and in collaboration with Dr.
John Buynak.
Click here to view structure or download coordinates.
(click on Publications to
find references).
We also
determined the crystal structure of the beta-lactamase KPC-2. KPC-2 has
special hydrolytic properties in that it can also hydrolyze carbapenems
and cephamycins which have a bulky alpha-substituent on the beta-lactam
ring. The ability to hydrolyze carbapenems is particularly worrisome
since carbapenem antibiotics constitute our 'last resort' antibiotics.
KPC-2 belongs therefore to the class of carbapenemases and its presence
has been linked to numerous difficult-to-treat Klebsiella outbreaks in
New York, Israel, and elsewhere. This rapid spread is further
facilitates since KPC-2 is the first plasmid-encoded class A
carbapenemase. Our structure elucidates the structural basis of the
carbapenemase activity and could aid in the development of more potent
novel beta-lactam antibiotics and inhibitors.
Our structure was recently
published in Biochemistry and is a collaboration with Dr. Bonomo (see
Publications link).
Click here to view structure or download coordinates.
We currently have a four-pronged approach of developing novel potent beta-lactamases inhibitors as we
explore: 1) C6-substituted penicillin sulfones (Nottingham et al. Bioorg & Med Chem. Lett 2011), 2) 6-alkylidene-2'-substituted penicillanic acid
sulfones (Bou et al., JACS 2010; Pattanaik et al. JBC 2009), 3) penam sulfones (Padayatti et al., JACS 2006; Sampson et al., Antimicrob. Agents and Chemotherap. 2011),
and 4) boronic acid transition state analogs (Ke et al., Antimicrob Agents and Chemotherap. 2011). These classes of compounds
are developed to target the most clinically urgent class A, class C, and class D beta-lactamases and carbapenemases.
Click here to visit
our Cleveland's joint VA/Case/CSU
ß-lactamase research group
Please click here to visit
Dr.Robert Bonomo's website http://id.clevelandactu.org/RBonomoCV.html
Please click here to visit Dr.
Marion Helfand's website http://www.case.edu/med/biochemistry/faculty/skalweit.html
Please click here to visit Dr.
Paul Carey's website http://www.cwru.edu/med/biochemistry/faculty/carey.html
Please click here to visit Dr.
John Buynak's website http://faculty.smu.edu/jbuynak/
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