We have developed techniques for the detailed analysis of cis-acting sequences in the pre-rRNA of Saccharomyces cerevisiae and used these to study the processing of internal transcribed spacer 1 (ITS1) leading to the synthesis of 5.8S rRNA. As is the case for many eukaryotes, the 5' end of yeast 5.8S rRNA is heterogeneous; we designate the major, short form 5.8S(S), and the minor form (which is seven or eight nucleotides longer) 5.8S(L). These RNAs do not have a precursor/product relationship, but result from the use of alternative processing pathways. In the major pathway, a previously unidentified processing site in ITS1, designated A3, is cleaved. A 10 nucleotide deletion at site A3 strongly inhibits processing of A3 and the synthesis of 5.8S(S); processing is predominantly transferred to the alternative 5.8S(L) pathway. Site A3 lies 76 nucleotides 5' to the end of 5.8S(S), and acts as an entry site for 5'-3' exonuclease digestion which generates the 5' end of 5.8S(S). This pathway is inhibited in strains mutant for XRNlp and RATlp. Both of these proteins have been reported to have 5'-3' exonuclease activity in vitro. Formation of 5.8S(L) is increased by mutations at A3 in cis or in RATlp and XRNlp in trans, and is kinetically faster than 5.8S(S) synthesis.

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The Interaction between carbon monoxide and the CO dehydrogenase from Clostidum thernoaceticum was studied by electron spin resonance (ESR) techniques. When the enzyme reacts with CO, a paramagnetic complex is formed which previously was shown, by Isotope substitution, to be due to a nickel-carbon species. In this paper, we demonstrate that iron is also a component of this ESR-detectable complex. When the iron in the enzyme is replaced with 57Fe, a broadening of 18 G in the g1I and 7 G in the go region is seen.This hyperfine interaction is probably due to more than one iron atom in the complex. Coenzyme A influences this ESR spectrum. In the absence of CoA, the ESR spectrum consists of two superimposed signals, which were simulated using the following ESR parameters: signal 1, with g = 2.074 and g = 2.028, and signal 2 with g, = 2.062, gy = 2.047, andg = 2.028.CoA converts signal 2 into signal 1. Since iron, nickel, and carbon all are part of this ESR-detectable complex, we propose that these atoms exist in a spin-coupled complex with net spin = 1/2, analogous to other iron-sulfur centers in which the metals are bridged by acid-labile sulfide.Carbon monoxide dehydrogenase is present in high concentration in the acetogenic bacteria (1, 2 Clearly the ESR spectrum of the paramagnetic CO dehydrogenase-C1 complex is due to nickel and carbon since hyperfine splitting was detected upon isotopic substitution with 61Ni (I = 3/2) and also when 13CO (I = 1/2) reacted with the enzyme (9). The major effects of the isotope substitutions were a splitting of the g = 2.028 (g I) component by "3CO and a broadening ofthe 2.074 (g1) component by the 61Ni nucleus.

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