Structure of the mammalian catalytic subunit of cAMP-dependent protein kinase and an inhibitor peptide displays an open conformation

Karlsson, Rolf; Zheng, Jie; Xuong, Nguyen-huu; Taylor, Susan S.; Sowadski, Janusz M.

doi:10.1107/s0907444993002306

Cited by 65 publications

(97 citation statements)

References 15 publications

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“…Whereas the large lobe (corresponding to residues 128-300) shows few changes, the small lobe (residues 40-127) rotates a substantial distance. A preliminary report of this binary structure, solved by an independent method, described the displacement of this small lobe (Karlsson et al, 1993). This concerted shift of the small lobe leads to an opening of the cleft.…”

Section: Comparison Of Models From the Mammalian And Recombinant Catamentioning

confidence: 98%

“…The nature of this motion is described by Karlsson et al (1993). This rotation alters the disposition of several conserved residues at the active site.…”

Section: Con Formational Changes In the Active Sitementioning

confidence: 99%

“…The opening of the active site is accomplished by a rotation of the antiparallel @-sheet through 15" (Karlsson et al, 1993). In understanding this movement it is also important to consider the portions of the molecule that do not change.…”

Section: Rigid Features Of the Small Lobementioning

confidence: 99%

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Crystal structures of the myristylated catalytic subunit of cAMP‐dependent protein kinase reveal open and closed conformations

et al. 1993

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Three crystal structures, representing two distinct conformational states, of the mammalian catalytic subunit of CAMP-dependent protein kinase were solved using molecular replacement methods starting from the refined structure of the recombinant catalytic subunit ternary complex (Zheng, J., et al., 1993a, Biochemistry 32,2154-2161). These structures correspond to the free apoenzyme, a binary complex with an iodinated inhibitor peptide, and a ternary complex with both ATP and the unmodified inhibitor peptide. The apoenzyme and the binary complex crystallized in an open conformation, whereas the ternary complex crystallized in a closed conformation similar to the ternary complex of the recombinant enzyme. The model of the binary complex, refined at 2.9 A resolution, shows the conformational changes associated with the open conformation. These can be described by a rotation of the small lobe and a displacement of the C-terminal30 residues. This rotation of the small lobe alters the cleft interface in the active-site region surrounding the glycine-rich loop and Thr 197, a critical phosphorylation site. In addition to the conformational changes, the myristylation site, absent in the recombinant enzyme, was clearly defined in the binary complex. The myristic acid binds in a deep hydrophobic pocket formed by four segments of the protein that are widely dispersed in the linear sequence. The N-terminal40 residues that lie outside the conserved catalytic core are anchored by the N-terminal myristylate plus an amphipathic helix that spans both lobes and is capped by Trp 30. Both posttranslational modifications, phosphorylation and myristylation, contribute directly to the stable structure of this enzyme.

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Section: Comparison Of Models From the Mammalian And Recombinant Catamentioning

confidence: 98%

“…The nature of this motion is described by Karlsson et al (1993). This rotation alters the disposition of several conserved residues at the active site.…”

Section: Con Formational Changes In the Active Sitementioning

confidence: 99%

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Crystal structures of the myristylated catalytic subunit of cAMP‐dependent protein kinase reveal open and closed conformations

et al. 1993

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“…The hinging movements of PKA that cause the opening occur primarily within the dipeptide segment of Gly 125 and Gly 126 positioned between the ATP binding residues and helix D, as defined originally by Olah and co-workers (34). Rotations of helix C can be taken as a measure of the opening of PKA structures, and ranks structures from the most closed in structure 1CDK through other open conformations of PKA such as staurosporine bound (1STC (10), or the apoenzyme structures (1CTP (35,36) and 1J3H (37)) to the most open conformation of BIM2MolB from this study (Table III and ith respect to 1CDK. The greatest rotations occur in the structures 1J3H-molecule A (14.2°) and BIM2MolB (14.6°) (Table III).…”

Section: Figmentioning

confidence: 99%

The Protein Kinase C Inhibitor Bisindolyl Maleimide 2 Binds with Reversed Orientations to Different Conformations of Protein Kinase A

Gaßel

Breitenlechner

König

et al. 2004

Journal of Biological Chemistry

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As the key mediators of eukaryotic signal transduction, the protein kinases often cause disease, and in particular cancer, when disregulated. Appropriately selective protein kinase inhibitors are sought after as research tools and as therapeutic drugs; several have already proven valuable in clinical use. The AGC subfamily protein kinase C (PKC) was identified early as a cause of cancer, leading to the discovery of a variety of PKC inhibitors. Despite its importance and early discovery, no crystal structure for PKC has yet been reported. Therefore, we have co-crystallized PKC inhibitor bisindolyl maleimide 2 (BIM2) with PKA variants to study its binding interactions.

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“…31 The median of these two distances measured in the dynamic snapshots of the wild-type and the E230Q mutant (data not shown) are comparable to the corresponding distances in the open conformation crystal structure of 1CTP. 32,33 When these two distances are compared to the number of best-docked PKI conformations in the corresponding snapshots, we do not see obvious correlation between the fluctuation of the distances and the number of best-docked PKIs. These results are consistent with the finding using the hinge-bending angle, suggesting that PKI docking is not much affected by the dynamic fluctua-FIGURE 2 P-2 Arg of PKI interacts directly with (a) Glu170 and Glu230 in 1ATP, (b) Asp241 in the second period of E230Q mutant MD simulation.…”

Section: Structural Changes Due To E230q Mutationmentioning

confidence: 99%

E230Q mutation of the catalytic subunit of cAMP‐dependent protein kinase affects local structure and the binding of peptide inhibitor

Ung

McCammon

2006

Biopolymers

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The active site of the mammalian cAMP-dependent protein kinase catalytic subunit (C-subunit) has a cluster of nonconserved acidic residues-Glu127, Glu170, Glu203, Glu230, and Asp241-that are crucial for substrate recognition and binding. Studies have shown that the Glu230 to Gln mutant (E230Q) of the enzyme has physical properties similar to the wild-type enzyme and has decreased affinity for a short peptide substrate, Kemptide. However, recent experiments intended to crystallize ternary complex of the E230Q mutant with MgATP and protein kinase inhibitor (PKI) could only obtain crystals of the apo-enzyme of E230Q mutant. To deduce the possible mechanism that prevented ternary complex formation, we used the relaxed-complex method (Lin, J.-H., et al. J Am Chem Soc 2002, 24, 5632-5633) to study PKI binding to the E230Q mutant C-subunit. In the E230Q mutant, we observed local structural changes of the peptide binding site that correlated closely to the reduced PKI affinity. The structural changes occurred in the F-to-G helix loop and appeared to hinder PKI binding. Reduced electrostatic potential repulsion among Asp241 from the helix loop section and the other acidic residues in the peptide binding site appear to be responsible for the structural change. # 2005 Wiley Periodicals, Inc.

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Structure of the mammalian catalytic subunit of cAMP-dependent protein kinase and an inhibitor peptide displays an open conformation

Cited by 65 publications

References 15 publications

Crystal structures of the myristylated catalytic subunit of cAMP‐dependent protein kinase reveal open and closed conformations

Crystal structures of the myristylated catalytic subunit of cAMP‐dependent protein kinase reveal open and closed conformations

The Protein Kinase C Inhibitor Bisindolyl Maleimide 2 Binds with Reversed Orientations to Different Conformations of Protein Kinase A

E230Q mutation of the catalytic subunit of cAMP‐dependent protein kinase affects local structure and the binding of peptide inhibitor

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