Inactivation of a class A and a class C β-lactamase by 6β-(hydroxymethyl)penicillanic acid sulfone

Krisztina M. Papp-Wallace; Christopher R. Bethel; Thomas D. Gootz; Wenchi Shang; Justin Stroh; William Lau; Dale McLeod; Loren Price; Anthony Marfat; Anne Distler; Sarah M. Drawz; Hansong Chen; Emily Harry; Micheal Nottingham; Paul R. Carey; John D. Buynak; Robert A. Bonomo

doi:10.1016/j.bcp.2011.11.015

Inactivation of a class A and a class C β-lactamase by 6β-(hydroxymethyl)penicillanic acid sulfone

Krisztina M. Papp-Wallace, Christopher R. Bethel, Thomas D. Gootz, Wenchi Shang, Justin Stroh, William Lau, Dale McLeod, Loren Price, Anthony Marfat, Anne Distler, Sarah M. Drawz, Hansong Chen, Emily Harry, Micheal Nottingham, Paul R. Carey, John D. Buynak, Robert A. Bonomo

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18 Scopus citations

Abstract

β-Lactamase inhibitors (clavulanic acid, sulbactam, and tazobactam) contribute significantly to the longevity of the β-lactam antibiotics used to treat serious infections. In the quest to design more potent compounds and to understand the mechanism of action of known inhibitors, 6β-(hydroxymethyl) penicillanic acid sulfone (6β-HM-sulfone) was tested against isolates expressing the class A TEM-1 β-lactamase and a clinically important variant of the AmpC cephalosporinase of Pseudomonas aeruginosa, PDC-3. The addition of the 6β-HM-sulfone inhibitor to ampicillin was highly effective. 6β-HM-sulfone inhibited TEM-1 with an IC ₅₀ of 12 ± 2 nM and PDC-3 with an IC ₅₀ of 180 ± 36 nM, and displayed lower partition ratios than commercial inhibitors, with partition ratios (k _cat/k _inact) equal to 174 for TEM-1 and 4 for PDC-3. Measured for 20 h, 6β-HM-sulfone demonstrated rapid, first-order inactivation kinetics with the extent of inactivation being related to the concentration of inhibitor for both TEM-1 and PDC-3. Using mass spectrometry to gain insight into the intermediates of inactivation of this inhibitor, 6β-HM-sulfone was found to form a major adduct of +247 ± 5 Da with TEM-1 and +245 ± 5 Da with PDC-3, suggesting that the covalently bound, hydrolytically stabilized acyl-enzyme has lost a molecule of water (HOH). Minor adducts of +88 ± 5 Da with TEM-1 and +85 ± 5 Da with PDC-3 revealed that fragmentation of the covalent adduct can result but appeared to occur slowly with both enzymes. 6β-HM-sulfone is an effective and versatile β-lactamase inhibitor of representative class A and C enzymes.

Original language	English (US)
Pages (from-to)	462-471
Number of pages	10
Journal	Biochemical Pharmacology
Volume	83
Issue number	4
DOIs	https://doi.org/10.1016/j.bcp.2011.11.015
State	Published - Feb 15 2012
Externally published	Yes

Bibliographical note

Funding Information:
We thank Dr. Jean-Marie Frère (Université de Liège, Belgium) for the purified TEM-1 β-lactamase used in parts of this study. KMP-W is supported by the Veterans Affairs Career Development Program. PRC was supported by the National Institutes of Health ( R01 GM54072 ). JDB is supported by grant N-0871 from the Robert A. Welch Foundation . RAB is supported by the Veterans Affairs Merit Review Program (RAB), the National Institutes of Health ( RO1 AI063517-01 ), and Geriatric Research Education and Clinical Centre VISN 10 . Pfizer also supported this work.

Keywords

PDC-3
Sulfone
TEM-1
β-Lactam
β-Lactamase
β-Lactamase inhibitor

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Papp-Wallace, K. M., Bethel, C. R., Gootz, T. D., Shang, W., Stroh, J., Lau, W., McLeod, D., Price, L., Marfat, A., Distler, A., Drawz, S. M., Chen, H., Harry, E., Nottingham, M., Carey, P. R., Buynak, J. D., & Bonomo, R. A. (2012). Inactivation of a class A and a class C β-lactamase by 6β-(hydroxymethyl)penicillanic acid sulfone. Biochemical Pharmacology, 83(4), 462-471. https://doi.org/10.1016/j.bcp.2011.11.015

Papp-Wallace, KM, Bethel, CR, Gootz, TD, Shang, W, Stroh, J, Lau, W, McLeod, D, Price, L, Marfat, A, Distler, A, Drawz, SM, Chen, H, Harry, E, Nottingham, M, Carey, PR, Buynak, JD & Bonomo, RA 2012, 'Inactivation of a class A and a class C β-lactamase by 6β-(hydroxymethyl)penicillanic acid sulfone', Biochemical Pharmacology, vol. 83, no. 4, pp. 462-471. https://doi.org/10.1016/j.bcp.2011.11.015

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title = "Inactivation of a class A and a class C β-lactamase by 6β-(hydroxymethyl)penicillanic acid sulfone",

abstract = "β-Lactamase inhibitors (clavulanic acid, sulbactam, and tazobactam) contribute significantly to the longevity of the β-lactam antibiotics used to treat serious infections. In the quest to design more potent compounds and to understand the mechanism of action of known inhibitors, 6β-(hydroxymethyl) penicillanic acid sulfone (6β-HM-sulfone) was tested against isolates expressing the class A TEM-1 β-lactamase and a clinically important variant of the AmpC cephalosporinase of Pseudomonas aeruginosa, PDC-3. The addition of the 6β-HM-sulfone inhibitor to ampicillin was highly effective. 6β-HM-sulfone inhibited TEM-1 with an IC 50 of 12 ± 2 nM and PDC-3 with an IC 50 of 180 ± 36 nM, and displayed lower partition ratios than commercial inhibitors, with partition ratios (k cat/k inact) equal to 174 for TEM-1 and 4 for PDC-3. Measured for 20 h, 6β-HM-sulfone demonstrated rapid, first-order inactivation kinetics with the extent of inactivation being related to the concentration of inhibitor for both TEM-1 and PDC-3. Using mass spectrometry to gain insight into the intermediates of inactivation of this inhibitor, 6β-HM-sulfone was found to form a major adduct of +247 ± 5 Da with TEM-1 and +245 ± 5 Da with PDC-3, suggesting that the covalently bound, hydrolytically stabilized acyl-enzyme has lost a molecule of water (HOH). Minor adducts of +88 ± 5 Da with TEM-1 and +85 ± 5 Da with PDC-3 revealed that fragmentation of the covalent adduct can result but appeared to occur slowly with both enzymes. 6β-HM-sulfone is an effective and versatile β-lactamase inhibitor of representative class A and C enzymes.",

keywords = "PDC-3, Sulfone, TEM-1, β-Lactam, β-Lactamase, β-Lactamase inhibitor",

author = "Papp-Wallace, {Krisztina M.} and Bethel, {Christopher R.} and Gootz, {Thomas D.} and Wenchi Shang and Justin Stroh and William Lau and Dale McLeod and Loren Price and Anthony Marfat and Anne Distler and Drawz, {Sarah M.} and Hansong Chen and Emily Harry and Micheal Nottingham and Carey, {Paul R.} and Buynak, {John D.} and Bonomo, {Robert A.}",

note = "Funding Information: We thank Dr. Jean-Marie Fr{\`e}re (Universit{\'e} de Li{\`e}ge, Belgium) for the purified TEM-1 β-lactamase used in parts of this study. KMP-W is supported by the Veterans Affairs Career Development Program. PRC was supported by the National Institutes of Health ( R01 GM54072 ). JDB is supported by grant N-0871 from the Robert A. Welch Foundation . RAB is supported by the Veterans Affairs Merit Review Program (RAB), the National Institutes of Health ( RO1 AI063517-01 ), and Geriatric Research Education and Clinical Centre VISN 10 . Pfizer also supported this work. ",

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T1 - Inactivation of a class A and a class C β-lactamase by 6β-(hydroxymethyl)penicillanic acid sulfone

AU - Papp-Wallace, Krisztina M.

AU - Bethel, Christopher R.

AU - Gootz, Thomas D.

AU - Shang, Wenchi

AU - Stroh, Justin

AU - Lau, William

AU - McLeod, Dale

AU - Price, Loren

AU - Marfat, Anthony

AU - Distler, Anne

AU - Drawz, Sarah M.

AU - Chen, Hansong

AU - Harry, Emily

AU - Nottingham, Micheal

AU - Carey, Paul R.

AU - Buynak, John D.

AU - Bonomo, Robert A.

N1 - Funding Information: We thank Dr. Jean-Marie Frère (Université de Liège, Belgium) for the purified TEM-1 β-lactamase used in parts of this study. KMP-W is supported by the Veterans Affairs Career Development Program. PRC was supported by the National Institutes of Health ( R01 GM54072 ). JDB is supported by grant N-0871 from the Robert A. Welch Foundation . RAB is supported by the Veterans Affairs Merit Review Program (RAB), the National Institutes of Health ( RO1 AI063517-01 ), and Geriatric Research Education and Clinical Centre VISN 10 . Pfizer also supported this work.

PY - 2012/2/15

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N2 - β-Lactamase inhibitors (clavulanic acid, sulbactam, and tazobactam) contribute significantly to the longevity of the β-lactam antibiotics used to treat serious infections. In the quest to design more potent compounds and to understand the mechanism of action of known inhibitors, 6β-(hydroxymethyl) penicillanic acid sulfone (6β-HM-sulfone) was tested against isolates expressing the class A TEM-1 β-lactamase and a clinically important variant of the AmpC cephalosporinase of Pseudomonas aeruginosa, PDC-3. The addition of the 6β-HM-sulfone inhibitor to ampicillin was highly effective. 6β-HM-sulfone inhibited TEM-1 with an IC 50 of 12 ± 2 nM and PDC-3 with an IC 50 of 180 ± 36 nM, and displayed lower partition ratios than commercial inhibitors, with partition ratios (k cat/k inact) equal to 174 for TEM-1 and 4 for PDC-3. Measured for 20 h, 6β-HM-sulfone demonstrated rapid, first-order inactivation kinetics with the extent of inactivation being related to the concentration of inhibitor for both TEM-1 and PDC-3. Using mass spectrometry to gain insight into the intermediates of inactivation of this inhibitor, 6β-HM-sulfone was found to form a major adduct of +247 ± 5 Da with TEM-1 and +245 ± 5 Da with PDC-3, suggesting that the covalently bound, hydrolytically stabilized acyl-enzyme has lost a molecule of water (HOH). Minor adducts of +88 ± 5 Da with TEM-1 and +85 ± 5 Da with PDC-3 revealed that fragmentation of the covalent adduct can result but appeared to occur slowly with both enzymes. 6β-HM-sulfone is an effective and versatile β-lactamase inhibitor of representative class A and C enzymes.

AB - β-Lactamase inhibitors (clavulanic acid, sulbactam, and tazobactam) contribute significantly to the longevity of the β-lactam antibiotics used to treat serious infections. In the quest to design more potent compounds and to understand the mechanism of action of known inhibitors, 6β-(hydroxymethyl) penicillanic acid sulfone (6β-HM-sulfone) was tested against isolates expressing the class A TEM-1 β-lactamase and a clinically important variant of the AmpC cephalosporinase of Pseudomonas aeruginosa, PDC-3. The addition of the 6β-HM-sulfone inhibitor to ampicillin was highly effective. 6β-HM-sulfone inhibited TEM-1 with an IC 50 of 12 ± 2 nM and PDC-3 with an IC 50 of 180 ± 36 nM, and displayed lower partition ratios than commercial inhibitors, with partition ratios (k cat/k inact) equal to 174 for TEM-1 and 4 for PDC-3. Measured for 20 h, 6β-HM-sulfone demonstrated rapid, first-order inactivation kinetics with the extent of inactivation being related to the concentration of inhibitor for both TEM-1 and PDC-3. Using mass spectrometry to gain insight into the intermediates of inactivation of this inhibitor, 6β-HM-sulfone was found to form a major adduct of +247 ± 5 Da with TEM-1 and +245 ± 5 Da with PDC-3, suggesting that the covalently bound, hydrolytically stabilized acyl-enzyme has lost a molecule of water (HOH). Minor adducts of +88 ± 5 Da with TEM-1 and +85 ± 5 Da with PDC-3 revealed that fragmentation of the covalent adduct can result but appeared to occur slowly with both enzymes. 6β-HM-sulfone is an effective and versatile β-lactamase inhibitor of representative class A and C enzymes.

KW - PDC-3

KW - Sulfone

KW - TEM-1

KW - β-Lactam

KW - β-Lactamase

KW - β-Lactamase inhibitor

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ER -

Inactivation of a class A and a class C β-lactamase by 6β-(hydroxymethyl)penicillanic acid sulfone

Abstract

Bibliographical note

Keywords

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