DNA sequence of the mitochondrial hypervariable region II from the neandertal type specimen.

The DNA sequence of the second hypervariable region of the mitochondrial control region of the Neandertal type specimen, found in 1856 in central Europe, has been determined from 92 clones derived from eight overlapping amplifications performed from four independent extracts. When the reconstructed sequence is analyzed together with the previously determined DNA sequence from the first hypervariable region, the Neandertal mtDNA is found to fall outside a phylogenetic tree relating the mtDNAs of contemporary humans. The date of divergence between the mtDNAs of the Neandertal and contemporary humans is estimated to 465,000 years before the present, with confidence limits of 317,000 and 741,000 years. Taken together, the results support the concept that the Neandertal mtDNA evolved separately from that of modern humans for a substantial amount of time and lends no support to the idea that they contributed mtDNA to contemporary modern humans.

mtDNA analysis of Nile River Valley populations: A genetic corridor or a barrier to migration?

To assess the extent to which the Nile River Valley has been a corridor for human migrations between Egypt and sub-Saharan Africa, we analyzed mtDNA variation in 224 individuals from various locations along the river. Sequences of the first hypervariable segment (HV1) of the mtDNA control region and a polymorphic HpaI site at position 3592 allowed us to designate each mtDNA as being of "northern" or "southern" affiliation. Proportions of northern and southern mtDNA differed significantly between Egypt, Nubia, and the southern Sudan. At slowly evolving sites within HV1, northern-mtDNA diversity was highest in Egypt and lowest in the southern Sudan, and southern-mtDNA diversity was highest in the southern Sudan and lowest in Egypt, indicating that migrations had occurred bidirectionally along the Nile River Valley. Egypt and Nubia have low and similar amounts of divergence for both mtDNA types, which is consistent with historical evidence for long-term interactions between Egypt and Nubia. Spatial autocorrelation analysis demonstrates a smooth gradient of decreasing genetic similarity of mtDNA types as geographic distance between sampling localities increases, strongly suggesting gene flow along the Nile, with no evident barriers. We conclude that these migrations probably occurred within the past few hundred to few thousand years and that the migration from north to south was either earlier or lesser in the extent of gene flow than the migration from south to north.

A hominoid-specific nuclear insertion of the mitochondrial D-loop: implications for reconstructing ancestral mitochondrial sequences.

A nuclear integration of a mitochondrial control region sequence on human chromosome 9 has been isolated. PCR analyses with primers specific for the respective insertion-flanking nuclear regions showed that the insertion took place on the lineage leading to Hominoidea (gibbon, orangutan, gorilla, chimpanzee, and human) after the Old World monkey-Hominoidea split. The sequences of the control region integrations were determined for humans, chimpanzees, gorillas, orangutans, and siamangs. These sequences were then used to construct phylogenetic trees with different methods, relating them with several hominoid, Old Work monkey, and New World monkey mitochondrial control region sequences. Applying maximum-likelihood, neighbor-joining, and parsimony algorithms, the insertion clade was attached to the branch leading to the hominoid mitochondrial sequences as expected from the PCR-determined presence/absence of this integration. An unexpected long branch leading to the internal node that connects all insertion sequences was observed for the different phylogeny reconstruction procedures. This finding is not totally compatible with the lower evolutionary rate in the nucleus than in the mitochondrial compartment. We determined the unambiguous substitutions on the branch leading to the most recent common ancestor (MRCA) of the mitochondrial inserts according to the parsimony criterium. We propose that they are unlikely to have been caused by damage of the transposing nucleic acid and that they are probably due to a change in the evolutionary mode after the transposition.

A nuclear 'fossil' of the mitochondrial D-loop and the origin of modern humans.

Mammalian mitochondrial DNA sequences evolve more rapidly than nuclear sequences. Although the rapid rate of evolution is an advantage for the study of closely related species and populations, it presents a problem in situations where related species, used as outgroups in phylogenetic analyses, have accumulated so much change that multiple substitutions obliterate the phylogenetic information. However, mitochondrial DNA sequences are frequently inserted into the nuclear genome, where they presumably evolve as nuclear pseudogene sequences and therefore more slowly than their mitochondrial counterparts. Such sequences thus represent molecular 'fossils' that could shed light on the evolution of the mitochondrial genome and could be used as outgroups in situations where no appropriate outgroup species exist. Here we show that human chromosome 11 carries a recent integration of the mitochondrial control region that can be used to gain further insight into the origin of the human mitochondrial gene pool.

Hemocyanins in spiders, XXIII. Complete amino-acid sequence of subunit a of Eurypelma californicum hemocyanin.

The complete amino-acid sequence of subunit a of the hemocyanin of the tarantula Eurypelma californicum was determined by manual sequencing. By limited chymotrypsinolysis, subunit a is split into two fragments of 25 kDa and 40 kDa, respectively, only one single peptide bond being attacked. The whole chain contains 15 methionine residues, after cyanogen bromide cleavage, 15 peptides were identified indicating that one residue (Met85) was not split by the cyanogen bromide reaction. For subcleavages, trypsin, chymotrypsin, Staphylococcus aureus proteinase, and Astacus fluviatilis proteinase were employed. The total chain length comprises 627 amino-acid residues, carbohydrate side chains were not found.

Hemocyanins in spiders, XIX. Complete amino-acid sequence of subunit d from Eurypelma californicum hemocyanin, and comparison to chain e.

The complete primary structure of subunit d of the hemocyanin from the tarantula Eurypelma californicum was determined by manual micro sequencing. Subunit d of Mr = 73000 is split about in the middle of the chain during limited trypsinolysis, only one single bond being attacked. The whole chain contains 14 methionine residues and after cyanogen bromide cleavage 15 peptides could be isolated by gel and ion exchange chromatography and high pressure liquid chromatography. The cyanogen bromide peptides and the large (Mr = 34000 and 37000, respectively) fragments resulting from limited trypsinolysis, were further cleaved with trypsin, chymotrypsin, Staphylococcus aureus proteinase, formic acid, and Astacus fluviatilis proteinase, the latter being very useful in obtaining certain overlapping peptides. The total chain length is 627 residues. Carbohydrate side chains were not found. The sequence is discussed with respect to the gross physical properties of the subunit, to homologies with subunit e and the cleavage specifities of the enzymes employed.