Transphosphorylation of E-coli Proteins and Kinase Specificity
Reporter: Larry H Bernstein, MD, FCAP
http://pharmaceuticalintelligence.com/2013/03/04/transphosphory…i-ptoteins-and/
X Wu, Man-Ho Oh, HS Kim, D Schwartz,… Huber SC.
Front. Plant Sci. 3:262. http://dx.doi.org/ 10.3389/fpls.2012.00262
Four lines of evidence suggest that transphosphorylation of E. coli proteins by BRI1
is specific and therefore provides meaningful results:
(1) phosphorylation is
- not correlated with bacterial protein abundance;
(2) phosphosite stoichiometry, estimated by spectral counting, is also
- not related to protein abundance;
(3) a transphosphorylation motif emerged
- with strong preference for basic residues both N- and C-terminal to the phosphosites; and
(4) other protein kinases (BAK1, PEPR1, FLS2, and CDPKβ) phosphorylated a distinct set of E. coli proteins and phosphosites. http://fpls.com/Transphosphorylation_of_E._coli_proteins_during_production_of_recombinant_protein_kinases_provides_a_robust_system_to_characterize_kinase_specificity/
English: Structure of the BAK1 protein. Based on PyMOL rendering of PDB 2ims. (Photo credit: Wikipedia)
Escherichia coli – Scanning Electron Microscopy (Photo credit: Wikipedia)
English: Protein kinase CK2, a phosphorylation enzyme (Photo credit: Wikipedia)
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This is very insightful. There is no doubt that there is the bias you refer to. 42 years ago, when I was postdocing in biochemistry/enzymology before completing my residency in pathology, I knew that there were very influential mambers of the faculty, who also had large programs, and attracted exceptional students. My mentor, it was said (although he was a great writer), could draft a project on toilet paper and call the NIH. It can’t be true, but it was a time in our history preceding a great explosion. It is bizarre for me to read now about eNOS and iNOS, and about CaMKII-á, â, ã, ä – isoenzymes. They were overlooked during the search for the genome, so intermediary metabolism took a back seat. But the work on protein conformation, and on the mechanism of action of enzymes and ligand and coenzyme was just out there, and became more important with the research on signaling pathways. The work on the mechanism of pyridine nucleotide isoenzymes preceded the work by Burton Sobel on the MB isoenzyme in heart. The Vietnam War cut into the funding, and it has actually declined linearly since.
A few years later, I was an Associate Professor at a new Medical School and I submitted a proposal that was reviewed by the Chairman of Pharmacology, who was a former Director of NSF. He thought it was good enough. I was a pathologist and it went to a Biochemistry Review Committee. It was approved, but not funded. The verdict was that I would not be able to carry out the studies needed, and they would have approached it differently. A thousand young investigators are out there now with similar letters. I was told that the Department Chairmen have to build up their faculty. It’s harder now than then. So I filed for and received 3 patents based on my work at the suggestion of my brother-in-law. When I took it to Boehringer-Mannheim, they were actually clueless.
This is very insightful. There is no doubt that there is the bias you refer to. 42 years ago, when I was postdocing in biochemistry/enzymology before completing my residency in pathology, I knew that there were very influential mambers of the faculty, who also had large programs, and attracted exceptional students. My mentor, it was said (although he was a great writer), could draft a project on toilet paper and call the NIH. It can’t be true, but it was a time in our history preceding a great explosion. It is bizarre for me to read now about eNOS and iNOS, and about CaMKII-á, â, ã, ä – isoenzymes. They were overlooked during the search for the genome, so intermediary metabolism took a back seat. But the work on protein conformation, and on the mechanism of action of enzymes and ligand and coenzyme was just out there, and became more important with the research on signaling pathways. The work on the mechanism of pyridine nucleotide isoenzymes preceded the work by Burton Sobel on the MB isoenzyme in heart. The Vietnam War cut into the funding, and it has actually declined linearly since.
A few years later, I was an Associate Professor at a new Medical School and I submitted a proposal that was reviewed by the Chairman of Pharmacology, who was a former Director of NSF. He thought it was good enough. I was a pathologist and it went to a Biochemistry Review Committee. It was approved, but not funded. The verdict was that I would not be able to carry out the studies needed, and they would have approached it differently. A thousand young investigators are out there now with similar letters. I was told that the Department Chairmen have to build up their faculty. It’s harder now than then. So I filed for and received 3 patents based on my work at the suggestion of my brother-in-law. When I took it to Boehringer-Mannheim, they were actually clueless.