Structure, Function and in vitro Reconstitution of Escherichia coli Ribosomes

Author(s):  
Peter Traub
mBio ◽  
2014 ◽  
Vol 5 (3) ◽  
Author(s):  
Christopher W. Lennon ◽  
Kimberly C. Lemmer ◽  
Jessica L. Irons ◽  
Max I. Sellman ◽  
Timothy J. Donohue ◽  
...  

ABSTRACTDksA is a global regulatory protein that, together with the alarmone ppGpp, is required for the “stringent response” to nutrient starvation in the gammaproteobacteriumEscherichia coliand for more moderate shifts between growth conditions. DksA modulates the expression of hundreds of genes, directly or indirectly. Mutants lacking a DksA homolog exhibit pleiotropic phenotypes in other gammaproteobacteria as well. Here we analyzed the DksA homolog RSP2654 in the more distantly relatedRhodobacter sphaeroides, an alphaproteobacterium. RSP2654 is 42% identical and similar in length toE. coliDksA but lacks the Zn finger motif of theE. coliDksA globular domain. Deletion of the RSP2654 gene results in defects in photosynthetic growth, impaired utilization of amino acids, and an increase in fatty acid content. RSP2654 complements the growth and regulatory defects of anE. colistrain lacking thedksAgene and modulates transcriptionin vitrowithE. coliRNA polymerase (RNAP) similarly toE. coliDksA. RSP2654 reduces RNAP-promoter complex stabilityin vitrowith RNAPs fromE. coliorR. sphaeroides, alone and synergistically with ppGpp, suggesting that even though it has limited sequence identity toE. coliDksA (DksAEc), it functions in a mechanistically similar manner. We therefore designate the RSP2654 protein DksARsp. Our work suggests that DksARsphas distinct and important physiological roles in alphaproteobacteria and will be useful for understanding structure-function relationships in DksA and the mechanism of synergy between DksA and ppGpp.IMPORTANCEThe role of DksA has been analyzed primarily in the gammaproteobacteria, in which it is best understood for its role in control of the synthesis of the translation apparatus and amino acid biosynthesis. Our work suggests that DksA plays distinct and important physiological roles in alphaproteobacteria, including the control of photosynthesis inRhodobacter sphaeroides. The study of DksARsp, should be useful for understanding structure-function relationships in the protein, including those that play a role in the little-understood synergy between DksA and ppGpp.


Author(s):  
Bożena Bruhn-Olszewska ◽  
Vadim Molodtsov ◽  
Michał Sobala ◽  
Maciej Dylewski ◽  
Katsuhiko S. Murakami ◽  
...  

2020 ◽  
Vol 64 (2) ◽  
pp. 349-358
Author(s):  
Margaux R. Audett ◽  
Thomas J. Maresca

Abstract The kinetochore (KT) field has matured tremendously since Earnshaw first identified CENP-A, CENP-B, and CENP-C [1,2]. In the past 35 years, the accumulation of knowledge has included: defining the parts list, identifying epistatic networks of interdependence within the parts list, understanding the spatial organization of subcomplexes into a massive structure – hundreds of megadaltons in size, and dissecting the functions of the KT in its entirety as well as of its individual parts. Like nearly all cell and molecular biology fields, the structure–function paradigm has been foundational to advances in the KT field. A point nicely highlighted by the fact that we are at the precipice of the in vitro reconstitution of a functional KT holo complex. Yet conventional notions of structure cannot provide a complete picture of the KT especially since it contains an abundance of unstructured or intrinsically disordered constituents. The combination of structured and disordered proteins within the KT results in an assembled system that is functionally greater than the sum of its parts.


Author(s):  
Weiqing Han ◽  
Lei Li ◽  
Nicholas Pettit ◽  
Wen Yi ◽  
Robert Woodward ◽  
...  

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