The yrdC family of genes codes for proteins that occur both independently and as a domain in proteins that have been implicated in regulation. An example for the latter case is the sua5 gene from yeast. Sua5 was identified as a suppressor of a translation initiation defect in cytochrome c and is required for normal growth in yeast (Na JG, Pinto I, Hampsey M, 1992, Genetics 11:791–801). However, the function of the Sua5 protein remains unknown; Sua5 could act either at the transcriptional or the posttranscriptional levels to compensate for an aberrant translation start codon in the cyc gene. To potentially learn more about the function of YrdC and proteins featuring this domain, the crystal structure of the YrdC protein from Escherichia coli was determined at a resolution of 2.0 Å. YrdC adopts a new fold with no obvious similarity to those of other proteins with known three-dimensional (3D) structure. The protein features a large concave surface on one side that exhibits a positive electrostatic potential. The dimensions of this depression, its curvature, and the fact that conserved basic amino acids are located at its floor suggest that YrdC may be a nucleic acid binding protein. An investigation of YrdC's binding affinities for single- and double-stranded RNA and DNA fragments as well as tRNAs demonstrates that YrdC binds preferentially to double-stranded RNA. Our work provides evidence that 3D structures of functionally uncharacterized gene products with unique sequences can yield novel folds and functional insights.

The trematode Fasciola hepatica secretes a number of cathepsin L-like proteases that are proposed to be involved in feeding, migration, and immune evasion by the parasite. To date, six full cDNA sequences encoding cathepsin L preproproteins have been identified. Previous studies have demonstrated that one of these cathepsins (L2) is unusual in that it is able to cleave substrates with a proline in the P2 position, translating into an unusual ability (for a cysteine proteinase) to clot fibrinogen. In this study, we report the sequence of a novel cathepsin (L5) and compare the substrate specificity of a recombinant enzyme with that of recombinant cathepsin L2. Despite sharing 80% sequence identity with cathepsin L2, cathepsin L5 does not exhibit substantial catalytic activity against substrates containing proline in the P2 position. Molecular modeling studies suggested that a single amino acid change (L69Y) in the mature proteinases may account for the difference in specificity at the S2 subsite. Recombinant cathepsin L5/L69Y was expressed in yeast and a substantial increase in the ability of this variant to accommodate substrates with a proline residue in the P2 position was observed. Thus, we have identified a single amino acid substitution that can substantially influence the architecture of the S2 subsite of F. hepatica cathepsin L proteases.

The binding capacity of the L-leucine receptor from Escherichia coli was measured with L-phenylalanine and 4-fluoro-L-phenylalanine as substrates by fluorescence. The apparent dissociation constants (KD) for L-leucine, L-phenylalanine, and 4-fluoro-L-phenylalanine are 0.40, 0.18, and 0.26 respectively. 19F NMR data show protein-induced shifts for the 4-fluoro-L-phenylalanine peak and 3-fluoro-L-phenylalanine when receptor is present. Evidence points to the binding of only the L-isomers of these fluorine analogs.

The folding of ribonuclease A (RNase A) has been extensively studied by characterizing the disulfide containing intermediates using different experimental conditions and analytical techniques. So far, some aspects still remain unclear such as the role of the loop 65–72 in the folding pathway. We have studied the oxidative folding of a RNase A derivative containing at position 67 the substitution Asn → isoAsp where the local structure of the loop 65–72 has been modified keeping intact the C65–C72 disulfide bond. By comparing the folding behavior of this mutant to that of the wild-type protein, we found that the deamidation significantly decreases the folding rate and alters the folding pathway of RNase A. Results presented here shed light on the role of the 65–72 region in the folding process of RNase A and also clarifies the effect of the deamidation on the folding/unfolding processes. On a more general ground, this study represents the first characterization of the intermediates produced along the folding of a deamidated protein.

In the Note Added in Proof that was published in the above article (Volume 9, Number 10, 2000, pp. 1857–1865), the wrong masses of the dummy atom (DZ) and the zinc atom (ZN) were used. The correct ones are 3.0 and 53.38, respectively.

Please note the corrections to the above referenced paper that appeared in Volume 9, Number 11, 2000, pages 2109–2117. On page 2110 (line 22) the KA has been mislabeled as Kdiss (the KD). Also in the legend to Figure 2, KD should be replaced with the KA (on line 2 and in Equation 7 on page 2111).

We inadvertently neglected to request permission for Figure 7 (Volume 8, Number 6, 1999, pp. 1166–1180), but we have received permission. The original figure is reprinted from the book Transition States of Biochemical Processes. It actually comes from Chapter 3 (Figure 1) of the book. The complete reference is: “Quantum Mechanical Approaches to the Study of Enzymatic Transition States and Reaction Paths.” Gerald A. Maggiora and Ralph E. Christoffersen, Transition States of Biochemical Processes, Edited by Richard D. Gandour and Richard L. Schowen, Plenum Press, New York, 1978, pp. 119–163 (Chapter 3).

Please note the corrections to the above referenced paper that appeared in Volume 9, Number 11, 2000, pages 2109–2117. On page 2110 (line 22) the KA has been mislabeled as Kdiss (the KD). Also in the legend to Figure 2, KD should be replaced with the KA (on line 2 and in Equation 7 on page 2111).

In the Note Added in Proof that was published in the above article (Volume 9, Number 10, 2000, pp. 1857–1865), the wrong masses of the dummy atom (DZ) and the zinc atom (ZN) were used. The correct ones are 3.0 and 53.38, respectively.