BACKGROUND
originally discovered as a host factor required for replication of phage qβ rna  <cit> , hfq  has emerged as a multifunctional regulator with a variety of different targets in the bacterial cell.
it forms a highly stable hexameric ring-shaped structure of identical subunits  that can polymerize as helical fibers  <cit> .
as a member of the sm/lsm protein family that is involved in rna metabolism, hfq contains an n-terminal α-helical domain followed by an antiparallel five-stranded beta-sheet, which is the hallmark of the so-called sm domain  <cit> .
by its interaction with rna, it controls the expression of many genes, stimulates the degradation of various mrnas and modulates rna processing events  <cit> .
its chaperone activity plays an important role for interactions between regulatory small non-coding rnas  and their target messenger rnas  <cit> .
hfq preferentially binds to a/u- rich single stranded regions, especially to poly tails and u-rich elements close to hairpin structures, where it induces structural changes  <cit> .
while the major part of hfq is found in the cytoplasm, a certain amount binds to genomic dna and is involved in nucleoid formation  <cit> .
furthermore, hfq stimulates the addition of poly tails to mrnas, where experimental data indicate that it interacts with rna and possibly also with pap  <cit> .
as a member of the nucleotidyltransferase family, this polymerase is closely related to trna nucleotidyltransferase , an enzyme that catalyzes the addition of the cca terminus to trna 3'-ends  <cit> .
here, it is demonstrated that these two enzymes do not only share structural as well as functional elements  <cit> , but are also both modulated in their activity by hfq.
RESULTS
hfq stimulates cca-addition
to investigate whether hfq has an effect on the catalytic activity of the e. coli cca-adding enzyme, the recombinant enzyme was incubated at 30°c for up to  <dig> minutes in the presence of ntps and hfq.
as a radioactively labeled substrate for cca-addition, yeast trnaphe lacking the cca terminus was used.
this is one of the best characterized trna molecules, and the corresponding unmodified in vitro transcript has a three-dimensional structure very similar to that of the fully modified in vivo molecule  <cit> .
it is therefore an ideal substrate for in vitro trna processing and aminoacylation reactions  <cit> .
in control experiments, addition of hfq was omitted or replaced by bovine serum albumin , which is frequently used to stabilize purified enzymes in an active conformation.
furthermore, additional prokaryotic rna binding proteins   <cit> , e. coli rnase p protein  <cit>  and e. coli hu  <cit> ) as well as two variants of hfq were used as controls.
hfq k56a, located in the highly conserved cavity of the hexamer, interferes with binding of the small regulatory rna dsra  <cit> .
hfq v43r, on the other hand, affects binding of polyadenylated rpso mrna and stimulation of pap  <cit> .
the reaction products were separated on polyacrylamide gels and visualized by autoradiography.
as shown in fig.
1a, cca-addition leads to new trna species with reduced electrophoretic mobility, where the uppermost signal corresponds to the trna carrying a complete cca terminus, while the other products indicate partial cca-addition .
while this activity is found in all assays, the time course indicates that hfq enhances the reaction efficiency substantially.
after  <dig> minutes of incubation, a strong increase of the reaction product  can be seen in the reaction assay including hfq, while the presence of bsa leads only to a moderate product increase relative to the reaction catalyzed by the cca-adding enzyme alone.
at longer incubation times, the reactions follow this trend.
however, this stimulation is not the result of an hfq- or bsa-catalyzed nucleotide incorporation, since incubation of trna substrate with ntps and hfq or bsa alone does not lead to any detectable reaction product .
the results were verified in  <dig> independent experiments.
reproductions of this assay with other trna transcripts without cca end  further corroborated the general hfq-mediated enhancement of cca-addition .
the additional control experiments with rna binding proteins and hfq variants shown in fig.
1b support these findings.
none of the rna binding proteins shows a stimulation of cca-addition comparable to that of hfq, indicating that this enhancement is indeed a specific effect of this protein.
furthermore, the fact that both hfq variants stimulate cca-addition comparable to the wild type protein demonstrates that this effect is different from pap stimulation or interaction with small regulatory rnas, where these point mutations lead to reduced efficiencies  <cit> .
interaction of hfq with trna substrate
while hfq visibly stimulates the cca-addition, it was not clear whether this enhancement was based on an interaction of hfq with the cca-adding enzyme or with the trna substrate.
therefore, gel shift experiments with radioactively labeled trna and hfq were performed under conditions identical to those of the cca-addition.
<dig>  pmol hfq and  <dig> pmol 32p-labeled trna substrate without cca-end were incubated for  <dig> and  <dig> minutes.
subsequently, samples were separated on a non-denaturing polyacrylamide gel and visualized by autoradiography .
neither bsa nor the cca-adding enzyme  lead to a shifted signal position of the trna, indicating that these proteins do not bind stably to the trna substrate.
in contrast, hfq alone as well as in combination with cca-adding enzyme binds to trna and leads to a shifted migration position of the transcript.
again, other trna transcripts lacking the cca terminus  led to identical results, indicating that the observed binding of hfq is not restricted to one type of trna molecule .
in a competition experiment, a nonspecific in vitro run-off transcript corresponding to part of the pcr  <dig>  topo vector sequence could not compete for hfq binding at any tested concentration.
however, a 5- to 30-fold excess of a transcript representing the 3'-terminal sequence of the e. coli rpso mrna, a well characterized hfq interaction partner  <cit> , could efficiently replace the trna in the binding assay .
these data clearly indicate that the binding of hfq to the trna transcript is specific and does not represent an in vitro artifact.
therefore, it is likely that the hfq-trna interaction is the basis for an enhanced cca-addition in the presence of hfq, while the stimulatory effect of bsa is probably unspecific and based on stabilization of the active conformation of the enzyme .
the hfq-trna complex was also observed at varying concentrations of hfq and trna .
these ratios correspond to rna-hfq concentrations used in other gel shift experiments  <cit>  and indicate that the interaction with trna molecules is stable over a range of varying concentrations of rna and hfq.
interestingly, the observed interaction of hfq with trna is not influenced by the presence or absence of a complete or partial cca-terminus, since control gel shift experiments with trnaphe ending with c--, cc- or cca showed that all transcript versions were bound with similar affinities .
this is in good agreement with the fact that the primary task of the cca-adding enzyme in e. coli is the repair of such partial cca termini, indicating a function of the hfq-trna complex in the restoration of cca termini.
kinetic analysis
to investigate the stimulating effect of hfq on cca-addition in more detail, michaelis-menten kinetic analyses were performed .
in five independent experiments, apparent km and vmax values were determined in presence and absence of hfq or bsa, respectively.
concentration of trnaphe without cca end was ranging from  <dig>  μm to  <dig>  μm.
reaction products were separated by page, and the signal intensities were quantified.
while the presence of hfq did not change km values compared to reaction of the cca-adding enzyme alone , it increases the maximum reaction rate of cca-addition from  <dig>  nm/min to  <dig>  nm/min .
on the other hand, the stimulating effect of bsa had only a slight impact on vmax, but lowered km.
this stimulating effect can be observed particularly at very low substrate concentrations , where it seems to be equivalent to the hfq-based reaction enhancement.
at higher trna concentrations, however, bsa leads only to a slight increase in reaction velocity compared to the reaction in the presence of hfq .
discussion
hfq is a multifunctional regulator in rna metabolism of prokaryotes  <cit> .
one of its functions is to modulate rna polyadenylation catalyzed by pap  <cit> .
this enzyme is closely related to the cca-adding enzyme that synthesizes and maintains the 3'-terminal cca sequence on trnas.
due to sequence similarities, it is discussed that these enzymes share a common ancestor and interconverted into each other during evolution  <cit> .
this fact led to the question whether the activity of these related enzymes is also modulated by identical proteins.
since hfq has a well characterized stimulating effect on pap, it was therefore investigated whether it also influences the activity of the cca-adding enzyme.
here, it is shown that hfq is indeed stimulating the cca-adding enzyme from e. coli.
while it is well known that bsa has a non-specific stabilizing effect on enzyme preparations, leading also to reaction enhancement, the hfq effect cannot be explained by simple protein stabilization alone, since it significantly exceeds the influence of bsa.
other rna binding proteins are also much less efficient than hfq in stimulating cca-addition , which is a further indication that hfq exerts a specific effect.
hence, pap as well as cca-adding enzyme are both stimulated by this protein in their activities.
the stimulation of polyadenylation is based on interaction of hfq with the rna substrate, where it binds as a hexameric ring of identical subunits to a/u-rich single-stranded elements closely located to hairpin-like structures  <cit> .
this effect on polyadenylation was also observed in vivo, where hfq might also interact with pap  <cit> .
however, hfq binding is not restricted to rna transcripts carrying such structural recognition elements, but binds also to other, not yet identified sequences, as it was demonstrated for small regulatory rna molecules and trnas, which is in agreement with our results  <cit> .
in the case of trna precursor transcripts carrying leader and trailer sequences, hfq seems to regulate trna maturation by slowing down rnase e-catalyzed cleavage, as both proteins recognize similar elements  <cit> .
the trna transcripts, however, that were used as substrates for cca-addition obviously do not carry typical hfq binding motifs but belong to the pool of transcripts that interact with hfq even in the absence of those elements.
this trna – hfq recognition is obviously mediated by amino acid residues of the protein that differ from those identified for interactions with mrna or small regulatory rnas.
indeed, replacement of v <dig> by r only affects stimulation of polyadenylation and interaction with mrna  <cit> , while substitution of k <dig> by a interferes with ncrna stabilization  <cit> .
yet, both variants do not affect the enhancement of cca-addition.
interestingly, a strong hfq-specific stimulation in cca-addition was only observed at trna concentrations at  <dig>  μm and above, while at lower concentration, the presence of bsa led to a comparable enhancement.
since in this experiment the trna concentration was increased, while the hfq concentration remained constant, the hfq-trna ratio was dropping from 19: <dig> to  <dig> : <dig> .
this led to a significant increase of the reaction rate .
therefore, it is possible that at low trna concentrations, a higher number of hfq hexamers binds to the trna and blocks the interaction with the cca-adding enzyme, forming an inactive hfq/trna complex which might reduce the cca incorporation to some degree.
this interpretation is supported by the observation of supershifted bands  at increased concentrations of hfq in gel shift experiments with radioactively labeled trna, indicating the binding of further hfq molecules to the trna .
the presence of such an excess of hfq can interfere with enzyme – substrate interaction and is likely to lead to an increase of apparent km values.
such inhibiting hfq complexes were also observed with the small regulatory rna dsra, where stoichiometry of hfq interaction determines the translatability of rpos mrna  <cit> .
at higher substrate trna concentrations, the inhibitory effect disappeared, presumably because less hfq molecules bind to an individual trna, rendering it accessible for the cca-adding enzyme.
how hfq increases the reaction velocity but does not influence km is not clear yet.
apparently, it exerts an indirect effect on the cca-adding enzyme, since no supershift of trna, hfq and cca-adding enzyme was observed in the gel shift analysis , and the kinetic data indicate that the affinity of the cca-adding enzyme to the trna is not affected by hfq.
the absence of such a supershifting complex shows that although such an interaction has to occur during the enhanced cca-addition, it is not stable or persistent enough to be detected in the experiment.
however, it is conceivable and in agreement with the absence of a supershift with cca-adding enzyme that the hfq/trna complex facilitates product release by weakening hydrogen bonds or ionic interactions between enzyme and trna.
subsequently, hfq might also release the completed trna and cycle to a new substrate for cca-addition.
a similar dissociation of hfq and rna was already observed  <cit> .
taken together, it seems that hfq is involved in the cca-end restoration in e. coli, where about 15% of the trnas have a damaged 3'-end  <cit> .
the abundance of  <dig>  hfq hexamers per cell is likely to be sufficient to enhance the repair of these truncated cca-termini  <cit> .
additionally, there is growing evidence that hfq-related lsm-like proteins in other organisms are also participating in trna processing, as depletion of lsm-like proteins in yeast led to the accumulation of several trna precursors  <cit> .
in this case the lsm proteins obviously do not compete for trna binding , but seem to recruit other processing factors to enhance trna maturation.
nevertheless, these data indicate that lsm-like proteins  might have a general role in the processing pathway of trna transcripts.
CONCLUSIONS
the results reported here indicate that the multifunctional lsm-like protein hfq stimulates not only the activity of poly polymerase, but also the reaction catalyzed by the cca-adding enzyme, where it forms a complex with the trna substrate.
the formation of this complex between hfq and trna is particularly interesting, as it demonstrates a hitherto unknown specific and thermodynamically stable interaction of this protein with a mature trna transcript lacking the cca-terminus.
the kinetic analysis indicates that this complex does not affect the substrate affinity of the cca-adding enzyme, but probably enhances the release of the reaction product, which is consistent with the fact that hfq, trna and cca-adding enzyme do not form a stable tertiary complex in the gel shift experiments.
taken together, hfq can stimulate the activity of the two closely related nucleotidyltransferases poly polymerase and cca-adding enzyme.
it will be interesting to see whether – and how – this protein modulates also the activities of other members of this enzyme class.

 originally discovered as a host factor required for replication of phage qβ rna, hfq has emerged as a multifunctional regulator with a variety of different targets in the bacterial cell. it forms a highly stable hexameric ring-shaped structure of identical subunits that can polymerize as helical fibers. as a member of the sm/lsm protein family that is involved in rna metabolism, hfq contains an n-terminal α-helical domain followed by an antiparallel five-stranded beta-sheet, which is the hallmark of the so-called sm domain. by its interaction with rna, it controls the expression of many genes, stimulates the degradation of various mrnas and modulates rna processing events. its chaperone activity plays an important role for interactions between regulatory small non-coding rnas and their target messenger rnas. hfq preferentially binds to a/u- rich single stranded regions, especially to poly tails and u-rich elements close to hairpin structures, where it induces structural changes. while the major part of hfq is found in the cytoplasm, a certain amount binds to genomic dna and is involved in nucleoid formation. furthermore, hfq stimulates the addition of poly tails to mrnas, where experimental data indicate that it interacts with rna and possibly also with pap. as a member of the nucleotidyltransferase family, this polymerase is closely related to trna nucleotidyltransferase, an enzyme that catalyzes the addition of the cca terminus to trna 3'-ends. here, it is demonstrated that these two enzymes do not only share structural as well as functional elements, but are also both modulated in their activity by hfq.  hfq stimulates cca-addition to investigate whether hfq has an effect on the catalytic activity of the e. coli cca-adding enzyme, the recombinant enzyme was incubated at 30°c for up to minutes in the presence of ntps and hfq. as a radioactively labeled substrate for cca-addition, yeast trnaphe lacking the cca terminus was used. this is one of the best characterized trna molecules, and the corresponding unmodified in vitro transcript has a three-dimensional structure very similar to that of the fully modified in vivo molecule. it is therefore an ideal substrate for in vitro trna processing and aminoacylation reactions. in control experiments, addition of hfq was omitted or replaced by bovine serum albumin, which is frequently used to stabilize purified enzymes in an active conformation. furthermore, additional prokaryotic rna binding proteins, e. coli rnase p protein and e. coli hu ) as well as two variants of hfq were used as controls. hfq k56a, located in the highly conserved cavity of the hexamer, interferes with binding of the small regulatory rna dsra. hfq v43r, on the other hand, affects binding of polyadenylated rpso mrna and stimulation of pap. the reaction products were separated on polyacrylamide gels and visualized by autoradiography. as shown in fig. 1a, cca-addition leads to new trna species with reduced electrophoretic mobility, where the uppermost signal corresponds to the trna carrying a complete cca terminus, while the other products indicate partial cca-addition. while this activity is found in all assays, the time course indicates that hfq enhances the reaction efficiency substantially. after minutes of incubation, a strong increase of the reaction product can be seen in the reaction assay including hfq, while the presence of bsa leads only to a moderate product increase relative to the reaction catalyzed by the cca-adding enzyme alone. at longer incubation times, the reactions follow this trend. however, this stimulation is not the result of an hfq- or bsa-catalyzed nucleotide incorporation, since incubation of trna substrate with ntps and hfq or bsa alone does not lead to any detectable reaction product. the results were verified in independent experiments. reproductions of this assay with other trna transcripts without cca end further corroborated the general hfq-mediated enhancement of cca-addition. the additional control experiments with rna binding proteins and hfq variants shown in fig. 1b support these findings. none of the rna binding proteins shows a stimulation of cca-addition comparable to that of hfq, indicating that this enhancement is indeed a specific effect of this protein. furthermore, the fact that both hfq variants stimulate cca-addition comparable to the wild type protein demonstrates that this effect is different from pap stimulation or interaction with small regulatory rnas, where these point mutations lead to reduced efficiencies. interaction of hfq with trna substrate while hfq visibly stimulates the cca-addition, it was not clear whether this enhancement was based on an interaction of hfq with the cca-adding enzyme or with the trna substrate. therefore, gel shift experiments with radioactively labeled trna and hfq were performed under conditions identical to those of the cca-addition. pmol hfq and pmol 32p-labeled trna substrate without cca-end were incubated for and minutes. subsequently, samples were separated on a non-denaturing polyacrylamide gel and visualized by autoradiography. neither bsa nor the cca-adding enzyme lead to a shifted signal position of the trna, indicating that these proteins do not bind stably to the trna substrate. in contrast, hfq alone as well as in combination with cca-adding enzyme binds to trna and leads to a shifted migration position of the transcript. again, other trna transcripts lacking the cca terminus led to identical results, indicating that the observed binding of hfq is not restricted to one type of trna molecule. in a competition experiment, a nonspecific in vitro run-off transcript corresponding to part of the pcr topo vector sequence could not compete for hfq binding at any tested concentration. however, a 5- to 30-fold excess of a transcript representing the 3'-terminal sequence of the e. coli rpso mrna, a well characterized hfq interaction partner, could efficiently replace the trna in the binding assay. these data clearly indicate that the binding of hfq to the trna transcript is specific and does not represent an in vitro artifact. therefore, it is likely that the hfq-trna interaction is the basis for an enhanced cca-addition in the presence of hfq, while the stimulatory effect of bsa is probably unspecific and based on stabilization of the active conformation of the enzyme. the hfq-trna complex was also observed at varying concentrations of hfq and trna. these ratios correspond to rna-hfq concentrations used in other gel shift experiments and indicate that the interaction with trna molecules is stable over a range of varying concentrations of rna and hfq. interestingly, the observed interaction of hfq with trna is not influenced by the presence or absence of a complete or partial cca-terminus, since control gel shift experiments with trnaphe ending with c--, cc- or cca showed that all transcript versions were bound with similar affinities. this is in good agreement with the fact that the primary task of the cca-adding enzyme in e. coli is the repair of such partial cca termini, indicating a function of the hfq-trna complex in the restoration of cca termini. kinetic analysis to investigate the stimulating effect of hfq on cca-addition in more detail, michaelis-menten kinetic analyses were performed. in five independent experiments, apparent km and vmax values were determined in presence and absence of hfq or bsa, respectively. concentration of trnaphe without cca end was ranging from μm to μm. reaction products were separated by page, and the signal intensities were quantified. while the presence of hfq did not change km values compared to reaction of the cca-adding enzyme alone, it increases the maximum reaction rate of cca-addition from nm/min to nm/min. on the other hand, the stimulating effect of bsa had only a slight impact on vmax, but lowered km. this stimulating effect can be observed particularly at very low substrate concentrations, where it seems to be equivalent to the hfq-based reaction enhancement. at higher trna concentrations, however, bsa leads only to a slight increase in reaction velocity compared to the reaction in the presence of hfq. discussion hfq is a multifunctional regulator in rna metabolism of prokaryotes. one of its functions is to modulate rna polyadenylation catalyzed by pap. this enzyme is closely related to the cca-adding enzyme that synthesizes and maintains the 3'-terminal cca sequence on trnas. due to sequence similarities, it is discussed that these enzymes share a common ancestor and interconverted into each other during evolution. this fact led to the question whether the activity of these related enzymes is also modulated by identical proteins. since hfq has a well characterized stimulating effect on pap, it was therefore investigated whether it also influences the activity of the cca-adding enzyme. here, it is shown that hfq is indeed stimulating the cca-adding enzyme from e. coli. while it is well known that bsa has a non-specific stabilizing effect on enzyme preparations, leading also to reaction enhancement, the hfq effect cannot be explained by simple protein stabilization alone, since it significantly exceeds the influence of bsa. other rna binding proteins are also much less efficient than hfq in stimulating cca-addition, which is a further indication that hfq exerts a specific effect. hence, pap as well as cca-adding enzyme are both stimulated by this protein in their activities. the stimulation of polyadenylation is based on interaction of hfq with the rna substrate, where it binds as a hexameric ring of identical subunits to a/u-rich single-stranded elements closely located to hairpin-like structures. this effect on polyadenylation was also observed in vivo, where hfq might also interact with pap. however, hfq binding is not restricted to rna transcripts carrying such structural recognition elements, but binds also to other, not yet identified sequences, as it was demonstrated for small regulatory rna molecules and trnas, which is in agreement with our results. in the case of trna precursor transcripts carrying leader and trailer sequences, hfq seems to regulate trna maturation by slowing down rnase e-catalyzed cleavage, as both proteins recognize similar elements. the trna transcripts, however, that were used as substrates for cca-addition obviously do not carry typical hfq binding motifs but belong to the pool of transcripts that interact with hfq even in the absence of those elements. this trna – hfq recognition is obviously mediated by amino acid residues of the protein that differ from those identified for interactions with mrna or small regulatory rnas. indeed, replacement of v by r only affects stimulation of polyadenylation and interaction with mrna, while substitution of k by a interferes with ncrna stabilization. yet, both variants do not affect the enhancement of cca-addition. interestingly, a strong hfq-specific stimulation in cca-addition was only observed at trna concentrations at μm and above, while at lower concentration, the presence of bsa led to a comparable enhancement. since in this experiment the trna concentration was increased, while the hfq concentration remained constant, the hfq-trna ratio was dropping from 19: to:. this led to a significant increase of the reaction rate. therefore, it is possible that at low trna concentrations, a higher number of hfq hexamers binds to the trna and blocks the interaction with the cca-adding enzyme, forming an inactive hfq/trna complex which might reduce the cca incorporation to some degree. this interpretation is supported by the observation of supershifted bands at increased concentrations of hfq in gel shift experiments with radioactively labeled trna, indicating the binding of further hfq molecules to the trna. the presence of such an excess of hfq can interfere with enzyme – substrate interaction and is likely to lead to an increase of apparent km values. such inhibiting hfq complexes were also observed with the small regulatory rna dsra, where stoichiometry of hfq interaction determines the translatability of rpos mrna. at higher substrate trna concentrations, the inhibitory effect disappeared, presumably because less hfq molecules bind to an individual trna, rendering it accessible for the cca-adding enzyme. how hfq increases the reaction velocity but does not influence km is not clear yet. apparently, it exerts an indirect effect on the cca-adding enzyme, since no supershift of trna, hfq and cca-adding enzyme was observed in the gel shift analysis, and the kinetic data indicate that the affinity of the cca-adding enzyme to the trna is not affected by hfq. the absence of such a supershifting complex shows that although such an interaction has to occur during the enhanced cca-addition, it is not stable or persistent enough to be detected in the experiment. however, it is conceivable and in agreement with the absence of a supershift with cca-adding enzyme that the hfq/trna complex facilitates product release by weakening hydrogen bonds or ionic interactions between enzyme and trna. subsequently, hfq might also release the completed trna and cycle to a new substrate for cca-addition. a similar dissociation of hfq and rna was already observed. taken together, it seems that hfq is involved in the cca-end restoration in e. coli, where about 15% of the trnas have a damaged 3'-end. the abundance of hfq hexamers per cell is likely to be sufficient to enhance the repair of these truncated cca-termini. additionally, there is growing evidence that hfq-related lsm-like proteins in other organisms are also participating in trna processing, as depletion of lsm-like proteins in yeast led to the accumulation of several trna precursors. in this case the lsm proteins obviously do not compete for trna binding, but seem to recruit other processing factors to enhance trna maturation. nevertheless, these data indicate that lsm-like proteins might have a general role in the processing pathway of trna transcripts.  the results reported here indicate that the multifunctional lsm-like protein hfq stimulates not only the activity of poly polymerase, but also the reaction catalyzed by the cca-adding enzyme, where it forms a complex with the trna substrate. the formation of this complex between hfq and trna is particularly interesting, as it demonstrates a hitherto unknown specific and thermodynamically stable interaction of this protein with a mature trna transcript lacking the cca-terminus. the kinetic analysis indicates that this complex does not affect the substrate affinity of the cca-adding enzyme, but probably enhances the release of the reaction product, which is consistent with the fact that hfq, trna and cca-adding enzyme do not form a stable tertiary complex in the gel shift experiments. taken together, hfq can stimulate the activity of the two closely related nucleotidyltransferases poly polymerase and cca-adding enzyme. it will be interesting to see whether – and how – this protein modulates also the activities of other members of this enzyme class. 
the bacterial sm-like protein hfq is known as an important regulator involved in many reactions of rna metabolism. furthermore, hfq binds specifically to trna transcripts, which seems to be the prerequisite for the observed effect on cca-addition. a prominent function of hfq is the stimulation of rna polyadenylation catalyzed by e. coli poly polymerase i. as a member of the nucleotidyltransferase superfamily, this enzyme shares a high sequence similarity with an other representative of this family, the trna nucleotidyltransferase that synthesizes the 3'-terminal sequence c-c-a to all trnas. in case of the cca-adding enzyme, however, the presented data indicate that the complex between hfq and trna substrate might enhance the product release from the enzyme.
 the bacterial sm-like protein hfq is known as an important regulator involved in many reactions of rna metabolism. a prominent function of hfq is the stimulation of rna polyadenylation catalyzed by e. coli poly polymerase i. as a member of the nucleotidyltransferase superfamily, this enzyme shares a high sequence similarity with an other representative of this family, the trna nucleotidyltransferase that synthesizes the 3'-terminal sequence c-c-a to all trnas. therefore, it was assumed that hfq might not only influence the poly polymerase in its specific activity, but also other, similar enzymes like the cca-adding enzyme.  based on the close evolutionary relation of these two nucleotidyltransferases, it was tested whether hfq is a specific modulator acting exclusively on pap or whether it also influences the activity of the cca-adding enzyme. the obtained data indicate that the reaction catalyzed by this enzyme is substantially accelerated in the presence of hfq. furthermore, hfq binds specifically to trna transcripts, which seems to be the prerequisite for the observed effect on cca-addition.  the increase of the cca-addition in the presence of hfq suggests that this protein acts as a stimulating factor not only for pap, but also for the cca-adding enzyme. in both cases, hfq interacts with rna substrates, while a direct binding to the corresponding enzymes was not demonstrated up to now. so far, the basic principle of these stimulatory effects is not clear yet. in case of the cca-adding enzyme, however, the presented data indicate that the complex between hfq and trna substrate might enhance the product release from the enzyme. 