N-utilizing proteins (Nus) for
m a co
mplex involved in the regulation of rRNA biosynthesis inenteric bacteria by
modulating the efficiency of transcriptional ter
mination [Nodwell, J. R., and Greenblatt,J. (1993)
Cell 72, 261-268]. The protein NusE (identical to the protein S10 of the s
mall riboso
mal subunit)fro
m the pathogenic
mycobacteriu
m M. tuberculosis has been cloned and overexpressed in
Escherichiacoli. The pure protein has been characterized by circular dichrois
m, ultracentrifugation, NMR, and bindingto NusB. The near-ultraviolet circular dichrois
m spectru
m of this protein suggests that it has a
moderate(ca. 12-16%)
![](/i<font color=)
mages/gifchars/alpha.gif" BORDER=0>-helical content at 30
![](/i<font color=)
mages/entities/deg.gif">C. The protein undergoes cold denaturation, with a te
mperature of
maxi
mu
m stability near 40
![](/i<font color=)
mages/entities/deg.gif">C, i
mplying a substantial heat capacity difference between the folded andunfolded states. The sedi
mentation equilibriu
m and velocity data indicate that the protein is
mono
mericand expanded in solution. NMR spectroscopy shows that there is no significant tertiary structure, andconfir
ms the low secondary structure content at low te
mperatures. Further
more, there was evidence for
more structure at 30
![](/i<font color=)
mages/entities/deg.gif">C than at 10
![](/i<font color=)
mages/entities/deg.gif">C. Well-defined shifts in peaks in the HSQC spectru
m of
15N labeledNusE/NusB when the unlabeled counterpart was added at approxi
mately stoichio
metric concentrationsshowed the for
mation of a NusE-NusB co
mplex in the absence of RNA. The far-UV CD andultracentrifuge experi
ments, however, indicated relatively weak binding. Isother
mal titration calori
metryshowed the binding was weak and endother
mic at 15
![](/i<font color=)
mages/entities/deg.gif">C, with a total
![](/i<font color=)
mages/gifchars/Delta.gif" BORDER=0 >
H of
![](/i<font color=)
mages/entities/ge.gif">10 kcal/
mol. This weakbinding is consistent with a s
mall interaction interface and lack of large confor
mational rearrange
mentsin the predo
minantly unfolded NusE protein. The confor
mational flexibility of NusE
may be i
mportantfor its roles in both the riboso
me and antiter
mination co
mplexes.