Can I read Importance of the A-helix of the catalytic subunit of cAMP-dependent protein kinase for stability and for orienting subdomains at the cleft interface: A-helix of CAMP-dependent protein kinase on EtoBox?

Importance of the A-helix of the catalytic subunit of cAMP-dependent protein kinase for stability and for orienting subdomains at the cleft interface: A-helix of CAMP-dependent protein kinase by Friedrich W. Herberg; Bastian Zimmermann; Maria Mcglone; Susan S. Taylor is a Biochemistry, Genetics and Molecular Biology article available to read on EtoBox.

What is Importance of the A-helix of the catalytic subunit of cAMP-dependent protein kinase for stability and for orienting subdomains at the cleft interface: A-helix of CAMP-dependent protein kinase about?

All eukaryotic protein kinases share a conserved catalytic core. In the catalytic (C) subunit of cAMP-dependent protein kinase (cAPK) this core is preceded by a myristylation motif followed by a long helix with Trp 30 at the end of this A-helix filling a hydrophobic cavity between the two lobes of the core. To understand the importance of the A-helix, the myristylation motif (delta 1-14) as well as the entire N-terminal segment (delta 1 -39) were deleted. In addition, Trp 30 was replaced with both Tyr and Ala. All proteins were overexpressed in E. coli and purified to homogeneity. rC(delta 1-14), rC(W30Y), and rC(W30A) all had reduced thermostability, but were catalytically indistinguishable from wild-type C. Based on Surface Plasmon Resonance, all three also formed stable holoenzyme complexes with the RI-subunit, although the appKds were reduced by more than 10-fold due to decrease in the association rate. Surprisingly, however, the holoenzymes were even more thermostable than wild-type holoenzyme. To obtain active enzyme, it was necessary to purify rC(delta 1-39) as a fusion protein with glutathione-S-transferase (GST-rC(delta 1-39), although its thermostability (Tm) was decrease

Who reads Importance of the A-helix of the catalytic subunit of cAMP-dependent protein kinase for stability and for orienting subdomains at the cleft interface: A-helix of CAMP-dependent protein kinase?

Author

Friedrich W. Herberg; Bastian Zimmermann; Maria Mcglone; Susan S. Taylor

Publisher

Cold Spring Harbor Laboratory Press; Wiley (John Wiley & Sons); Wiley-Blackwell; Wiley (ISSN 0961-8368)

Published

1997

Language

EN

Field

Biochemistry, Genetics and Molecular Biology (Life Sciences)