Action at a Distance

Blobaum, Anna L.; Xu, Shu; Rowlinson, Scott W.; Duggan, Kelsey C.; Banerjee, Surajit; Kudalkar, Shalley N.; Birmingham, William R.; Ghebreselasie, Kebreab; Marnett, Lawrence J.

doi:10.1074/jbc.m114.635987

Cited by 30 publications

(18 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…It may be that some detergents exert an indirect effect on the cyclooxygenase channel opening that alters some structural or electrostatic interaction required for aspirin to access the cyclooxygenase channel. The altered inhibition time course of celecoxib may implicate the region around the COX-2 specific side pocket housing Arg-513 58 , although there is no precedent to suggest this residue is involved in aspirin inhibition.…”

Section: Discussionmentioning

confidence: 99%

Crystal Structure of Aspirin-Acetylated Human Cyclooxygenase-2: Insight into the Formation of Products with Reversed Stereochemistry

et al. 2016

View full text Add to dashboard Cite

Aspirin and other nonsterroidal anti-inflammatory drugs target the Cyclooxygenase enzymes (COX-1 and COX-2) to block the formation of prostaglandins. Aspirin is unique in that it covalently modifies each enzyme by acetylating Ser-530 within the cyclooxygenase active site. Acetylation of COX-1 leads to complete loss of activity, while acetylation of COX-2 results in the generation of the mono-oxygenated product 15(R)-hydroxyeicosatetraenoic acid (15R-HETE). Ser-530 has also been shown to influence the stereochemistry for oxygen addition into the prostaglandin product. We determined the crystal structures of S530T murine (mu) COX-2, aspirin-acetylated human (hu) COX-2, and huCOX-2 in complex with salicylate to 1.9Å, 2.0Å, and 2.4Å, respectively. The structures reveal that: 1) the acetylated Ser-530 completely blocks access to the hydrophobic groove; 2) the observed binding pose of salicylate is reflective of the enzyme-inhibitor complex prior to acetylation; and 3) the observed Thr-530 rotamer in the S530T muCOX-2 crystal structure does not impede access to the hydrophobic groove. Based on these structural observations, along with functional analysis of the S530T/G533V double mutant, we propose a working hypothesis for the generation of 15R-HETE by aspirin-acetylated COX-2. We also observe differential acetylation of COX-2 purified in various detergent systems and nanodiscs, indicating that detergent and lipid binding within the membrane-binding domain of the enzyme alters the rate of the acetylation reaction in vitro.

show abstract

Section: Discussionmentioning

confidence: 99%

Crystal Structure of Aspirin-Acetylated Human Cyclooxygenase-2: Insight into the Formation of Products with Reversed Stereochemistry

et al. 2016

View full text Add to dashboard Cite

show abstract

“…Curiously, ibuprofen inhibition of huCOX-1 becomes time-dependent in the presence of nonsubstrate FAs (52). Double-tryptophan murine COX-2 mutants at positions 89/90 or 89/119 enhance the time dependence of ibuprofen and mefenamic acid (65). (R)-Profens are substrate-selective inhibitors of 2-AG oxygenation by COX-2 (61).…”

Section: Time-independent Inhibitorsmentioning

confidence: 99%

Interactions of fatty acids, nonsteroidal anti-inflammatory drugs, and coxibs with the catalytic and allosteric subunits of cyclooxygenases-1 and -2

Smith

Malkowski

2019

Journal of Biological Chemistry

View full text Add to dashboard Cite

Edited by George M. CarmanProstaglandin endoperoxide H synthases-1 and -2, commonly called cyclooxygenases-1 and -2 (COX-1 and -2), catalyze the committed step in prostaglandin biosynthesis-the conversion of arachidonic acid to prostaglandin endoperoxide H 2 . Both COX isoforms are sequence homodimers that function as conformational heterodimers having allosteric (Eallo) and catalytic (Ecat) subunits. At least in the case of COX-2, the enzyme becomes folded into a stable Eallo/Ecat pair. Some COX inhibitors (i.e. nonsteroidal anti-inflammatory drugs and coxibs) and common fatty acids (FAs) modulate Ecat activity by binding Eallo. However, the interactions and outcomes often differ between isoforms. For example, naproxen directly and completely inhibits COX-1 by binding Ecat but indirectly and incompletely inhibits COX-2 by binding Eallo. Additionally, COX-1 is allosterically inhibited up to 50% by common FAs like palmitic acid, whereas COX-2 is allosterically activated 2-fold by palmitic acid. FA binding to Eallo also affects responses to COX inhibitors. Thus, COXs are physiologically and pharmacologically regulated by the FA tone of the milieu in which each operates-COX-1 in the endoplasmic reticulum and COX-2 in the Golgi apparatus. Cross-talk between Eallo and Ecat involves a loop in Eallo immediately downstream of Arg-120. Mutational studies suggest that allosteric modulation requires a direct interaction between the carboxyl group of allosteric effectors and Arg-120 of Eallo; however, structural studies show some allosterically active FAs positioned in COX-2 in a conformation lacking an interaction with Arg-120. Thus, many details about the biological consequences of COX allosterism and how ligand binding to Eallo modulates Ecat remain to be resolved. Figure 4. Model for allosteric interactions between monomers of the COX-2 homodimer caused by the binding of FAs.Ecat is the catalytic monomer to which heme is bound, whereas Eallo is the allosteric monomer that does not bind heme. A, binding of AA to Eallo and Ecat of huCOX-2 and subsequent oxygenation of AA. The K d value for Ecat is the K m for AA (8.4 M) in the absence of other FAs. The K d value for AA binding to Eallo was determined to be 0.26 M. 6 B, binding of AA to Eallo and Ecat in the presence of PA. We make the following assumptions: (a) that the K d value for binding of AA to the Ecat site is equal to the K m value in the presence of PA (i.e. Ͻ 3.5 M), and (b) that the K d value for PA binding to the Ecat site is high (Ն50 M) because PA does not inhibit AA oxygenation by huCOX-2 even at low concentrations of AA. We estimate that the K d values for the binding of PA and stearic acid to the Eallo site are both ϳ7.5 M. 5 Note that the K d value for PA binding to Eallo is 30-fold that for AA binding to Eallo so that at all relevant FA concentrations AA and PA will compete for binding to Eallo, and the effect of PA will be determined by the PA/AA ratio. This figure and the legend are modified from Ref. 51. JBC REVIEWS: Allosteric regulation of COX-2

show abstract

“…As is seen for indomethacin, the Ser-530 hydroxyl group of COX-2 forms a hydrogen bond with the carbonyl between the indole and phenyl rings of the indomethacin moiety of compounds 1 and 2 in their respective crystal complexes. Mutation of Ser-530 to alanine decreased the potency of compound 1 (IC 50 ϭ 2.5 M) while minimally affecting the potency of com- Prior work had demonstrated that mutation of Val-89 to tryptophan alters the kinetics of inhibition of many weak reversible inhibitors so that they behave as time-dependent, tight-binding inhibitors (28). Crystal structure data revealed that the Trp-89 residue fills a gap between helixes B and D of the MBD, changing the opening of the lobby from a C-shaped partial ring to a fully closed donut shape.…”

Section: Structural Determinants Of Cox-2 Inhibition Of Compounds 1 Amentioning

confidence: 99%

Fluorescent indomethacin-dansyl conjugates utilize the membrane-binding domain of cyclooxygenase-2 to block the opening to the active site

Uddin

Banerjee

et al. 2019

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Action at a Distance

Cited by 30 publications

References 30 publications

Crystal Structure of Aspirin-Acetylated Human Cyclooxygenase-2: Insight into the Formation of Products with Reversed Stereochemistry

Crystal Structure of Aspirin-Acetylated Human Cyclooxygenase-2: Insight into the Formation of Products with Reversed Stereochemistry

Interactions of fatty acids, nonsteroidal anti-inflammatory drugs, and coxibs with the catalytic and allosteric subunits of cyclooxygenases-1 and -2

Fluorescent indomethacin-dansyl conjugates utilize the membrane-binding domain of cyclooxygenase-2 to block the opening to the active site

Contact Info

Product

Resources

About