The genetic structure of the indigenous hunter-gatherer peoples of southern Africa, the oldest known lineage of modern human, is important for understanding human diversity. Studies based on mitochondrial1 and small sets of nuclear markers2 have shown that these hunter-gatherers, known as Khoisan, San, or Bushmen, are genetically divergent from other humans1,3. However, until now, fully sequenced human genomes have been limited to recently diverged populations4–8. Here we present the complete genome sequences of an indigenous hunter-gatherer from the Kalahari Desert and a Bantu from southern Africa, as well as protein-coding regions from an additional three hunter-gatherers from disparate regions of the Kalahari. We characterize the extent of whole-genome and exome diversity among the five men, reporting 1.3 million novel DNA differences genome-wide, including 13,146 novel amino acid variants. In terms of nucleotide substitutions, the Bushmen seem to be, on average, more different from each other than, for example, a European and an Asian. Observed genomic differences between the hunter-gatherers and others may help to pinpoint genetic adaptations to an agricultural lifestyle. Adding the described variants to current databases will facilitate inclusion of southern Africans in medical research efforts, particularly when family and medical histories can be correlated with genome-wide data.

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The thermally induced 3,3-sigmatropic shift of allyl vinyl ethers to γ,δ-unsaturated carbonyl compounds, i.e., the Claisen rearrangement (Scheme 1), is an important reaction in synthetic organic chemistry and is the prototype for the conversion of chorismate to prephenate in the biosynthesis of phenylalanine. 1 Because of the simplicity of the reaction, its mechanism has been the object of numerous experimental and theoretical studies. The concern of this Account is not only how the reaction proceeds but how well theory reproduces the experimentally observed kinetic isotope effects, solvent effects, and substituent effects. It is our contention that, despite great progress, theory is lacking, particularly in the area of solvent effects where hydrophobic acceleration is an important factor. Indeed, we argue that hydrophobic effects are critically important in the Claisen rearrangement and in many other reactions, including solvolysis of tert-alkyl chlorides. Further, the response of the Claisen rearrangement to substituents is, in our view, predictable, not on the basis of an "aromatic" transition state of fixed geometry but rather on the basis of a transition state of variable geometry whose structure and stability respond to the nature and placement of substituents.The Claisen rearrangement is a unimolecular process with activation parameters that suggest a cyclic transition state. 2 Stereochemical studies have revealed transfer of asymmetry from the double bonds to the newly formed σ bond in a manner that suggests not only a transition state with cyclic delocalization of the six electrons of the original π bonds and the C-O σ bond but a three-dimensional geometry which resembles a chairlike structure (Scheme 1). 3 This would be consistent with the conservation of orbital symmetry rules. 4 However, the observations could also be rationalized by formation of a chairlike oxacyclohexanediyl or zwitterion-like transition state or intermedi-

Recombinant hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp) was reported to possess terminal transferase (TNTase) activity, the ability to add nontemplated nucleotides to the 3 end of viral RNAs. However, this TNTase was later purported to be a cellular enzyme copurifying with the HCV RdRp. In this report, we present evidence that TNTase activity is an inherent function of HCV and bovine viral diarrhea virus RdRps highly purified from both prokaryotic and eukaryotic cells. A change of the highly conserved GDD catalytic motif in the HCV RdRp to GAA abolished both RNA synthesis and TNTase activity. Furthermore, the nucleotides added via this TNTase activity are strongly influenced by the sequence near the 3 terminus of the viral template RNA, perhaps accounting for the previous discrepant observations between RdRp preparations. Last, the RdRp TNTase activity was shown to restore the ability to direct initiation of RNA synthesis in vitro on an initiation-defective RNA substrate, thereby implicating this activity in maintaining the integrity of the viral genome termini.Replication of plus-strand RNA viruses requires a multisubunit enzyme, the replicase, which is composed of viral and cellular factors (6). Biochemical characterization of eukaryotic replicases is limited because of difficulty in obtaining sufficient quantities of purified replicase. Furthermore, the hepatitis C virus (HCV) replicase has not been reported to accept exogenously provided RNAs. These results have prompted studies of the recombinant HCV RNA-dependent RNA polymerase (RdRp), the subunit responsible for phosphoryl transfer (9,16,17,26,31,38). While RdRps lack many properties of replicases, they are useful for characterizing some fundamental activities, such as the recognition of the initiation site and the kinetics of nucleotide polymerization (4,18,24).The HCV RdRp has recently been demonstrated to initiate RNA synthesis preferentially from the 3Ј terminus of the template RNA (16,17,26,31). Initiation from the 3Ј terminus raises a potential problem that viruses might encounter: cellular RNases that degrade even a few 3Ј nucleotides could prevent the initiation of viral RNA replication. Several mechanisms have been proposed that might allow RNA viruses to preserve or restore the sequences at the termini of their genome. These include base-pairing-dependent and base-pairing-independent recombination (12), priming by oligonucleotides aborted during the initiation of RNA synthesis (29), telomerase-like addition of a repeated sequence (33), and nontemplated nucleotide addition (7,12). Also, terminal adenylyl transferase activity was found to be associated with poliovirus polymerase 3D pol (30), possibly causing restoration of infectivity of poliovirus RNAs lacking the wild-type poly(A) tail.Recombinant HCV RdRp was reported to possess the ability to add nontemplated nucleotides to the 3Ј end of viral RNAs (5). However, this terminal transferase (TNTase) activity was later purported to be a cellular enzyme copurifying with the HCV RdRp...

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