Role of second-largest RNA polymerase I subunit Zn-binding domain in enzyme assembly.
The second-largest subunits of eukaryal RNA polymerases are similar to the beta subunits of prokaryal RNA polymerases throughout much of their lengths. The second-largest subunits from eukaryal RNA polymerases contain a four-cysteine Zn-binding domain at their C termini. The domain is also present in archaeal homologs but is absent from prokaryal homologs. Here, we investigated the role of the C-terminal Zn-binding domain of Rpa135, the second-largest subunit of yeast RNA polymerase I. Analysis of nonfunctional Rpa135 mutants indicated that the Zn-binding domain is required for recruitment of the largest subunit, Rpa190, into the RNA polymerase I complex. Curiously, the essential function of the Rpa135 Zn-binding domain is not related to Zn(2+) binding per se, since replacement of only one of the four cysteine residues with alanine led to the loss of Rpa135 function. Even more strikingly, replacement of all four cysteines with alanines resulted in functional Rpa135.
Naryshkina
Tatyana
T
Waksman Institute, Rutgers, The State University, Piscataway, NJ 08854, USA.
Bruning
Adrian
A
Gadal
Olivier
O
Severinov
Konstantin
K
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002466:Cell Nucleolus / Q000737:chemistry; D005690:Galactose / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D012318:RNA Polymerase I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D014170:Transformation, Genetic; D015032:Zinc / Q000378:metabolism*
D021122:Protein Subunits; D012318:RNA Polymerase I; D005947:Glucose; D015032:Zinc; D005690:Galactose