PMID- 27150669
OWN - NLM
STAT- MEDLINE
DCOM- 20180209
LR  - 20181113
IS  - 2045-2322 (Electronic)
IS  - 2045-2322 (Linking)
VI  - 6
DP  - 2016 May 6
TI  - Mechanism of Ribonuclease III Catalytic Regulation by Serine Phosphorylation.
PG  - 25448
LID - 10.1038/srep25448 [doi]
LID - 25448
AB  - Ribonuclease III (RNase III) is a conserved, gene-regulatory bacterial 
      endonuclease that cleaves double-helical structures in diverse coding and 
      noncoding RNAs. RNase III is subject to multiple levels of control, reflective of 
      its global regulatory functions. Escherichia coli (Ec) RNase III catalytic 
      activity is known to increase during bacteriophage T7 infection, reflecting the 
      expression of the phage-encoded protein kinase, T7PK. However, the mechanism of 
      catalytic enhancement is unknown. This study shows that Ec-RNase III is 
      phosphorylated on serine in vitro by purified T7PK, and identifies the targets as 
      Ser33 and Ser34 in the N-terminal catalytic domain. Kinetic experiments reveal a 
      5-fold increase in kcat and a 1.4-fold decrease in Km following phosphorylation, 
      providing a 7.4-fold increase in catalytic efficiency. Phosphorylation does not 
      change the rate of substrate cleavage under single-turnover conditions, 
      indicating that phosphorylation enhances product release, which also is the 
      rate-limiting step in the steady-state. Molecular dynamics simulations provide a 
      mechanism for facilitated product release, in which the Ser33 phosphomonoester 
      forms a salt bridge with the Arg95 guanidinium group, thereby weakening RNase III 
      engagement of product. The simulations also show why glutamic acid substitution 
      at either serine does not confer enhancement, thus underscoring the specific 
      requirement for a phosphomonoester.
FAU - Gone, Swapna
AU  - Gone S
AD  - Department of Chemistry, Philadelphia PA, 19122, USA.
FAU - Alfonso-Prieto, Mercedes
AU  - Alfonso-Prieto M
AD  - Institute for Computational Molecular Science, Philadelphia PA, 19122, USA.
FAU - Paudyal, Samridhdi
AU  - Paudyal S
AD  - Department of Biology, Temple University, Philadelphia PA, 19122, USA.
FAU - Nicholson, Allen W
AU  - Nicholson AW
AD  - Department of Chemistry, Philadelphia PA, 19122, USA.
AD  - Department of Biology, Temple University, Philadelphia PA, 19122, USA.
LA  - eng
GR  - R01 GM056772/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
PT  - Research Support, Non-U.S. Gov't
DEP - 20160506
PL  - England
TA  - Sci Rep
JT  - Scientific reports
JID - 101563288
RN  - 0 (Escherichia coli Proteins)
RN  - 452VLY9402 (Serine)
RN  - 63231-63-0 (RNA)
RN  - EC 2.7.- (Protein Kinases)
RN  - EC 3.1.26.3 (Ribonuclease III)
RN  - EC 3.1.26.3 (ribonuclease III, E coli)
SB  - IM
MH  - Bacteriophage T7/enzymology
MH  - Escherichia coli/*enzymology
MH  - Escherichia coli Proteins/*metabolism
MH  - Hydrolysis
MH  - Kinetics
MH  - Molecular Dynamics Simulation
MH  - Phosphorylation
MH  - Protein Kinases/metabolism
MH  - *Protein Processing, Post-Translational
MH  - RNA/metabolism
MH  - Ribonuclease III/*metabolism
MH  - Serine/*metabolism
PMC - PMC4858673
EDAT- 2016/05/07 06:00
MHDA- 2018/02/10 06:00
PMCR- 2016/05/06
CRDT- 2016/05/07 06:00
PHST- 2016/01/08 00:00 [received]
PHST- 2016/04/15 00:00 [accepted]
PHST- 2016/05/07 06:00 [entrez]
PHST- 2016/05/07 06:00 [pubmed]
PHST- 2018/02/10 06:00 [medline]
PHST- 2016/05/06 00:00 [pmc-release]
AID - srep25448 [pii]
AID - 10.1038/srep25448 [doi]
PST - epublish
SO  - Sci Rep. 2016 May 6;6:25448. doi: 10.1038/srep25448.