SUMMARY Cellular processes often depend on stable physical associations between proteins. Despite recent progress, knowledge of the composition of human protein complexes remains limited. To close this gap, we applied an integrative global proteomic profiling approach, based on chromatographic separation of cultured human cell extracts into more than one thousand biochemical fractions which were subsequently analyzed by quantitative tandem mass spectrometry, to systematically identify a network of 13,993 high-confidence physical interactions among 3,006 stably-associated soluble human proteins. Most of the 622 putative protein complexes we report are linked to core biological processes, and encompass both candidate disease genes and unnanotated proteins to inform on mechanism. Strikingly, whereas larger multi-protein assemblies tend to be more extensively annotated and evolutionarily conserved, human protein complexes with 5 or fewer subunits are far more likely to be functionally un-annotated or restricted to vertebrates, suggesting more recent functional innovations.

Macromolecular complexes are essential to conserved biological processes, but their prevalence across animals is unclear. By combining extensive biochemical fractionation with quantitative mass spectrometry, we directly examined the composition of soluble multiprotein complexes among diverse metazoan models. Using an integrative approach, we then generated a draft conservation map consisting of >1 million putative high-confidence co-complex interactions for species with fully sequenced genomes that encompasses functional modules present broadly across all extant animals. Clustering revealed a spectrum of conservation, ranging from ancient Eukaryal assemblies likely serving cellular housekeeping roles for at least 1 billion years, ancestral complexes that have accrued contemporary components, and rarer metazoan innovations linked to multicellularity. We validated these projections by independent co-fractionation experiments in evolutionarily distant species, by affinity-purification and by functional analyses. The comprehensiveness, centrality and modularity of these reconstructed interactomes reflect their fundamental mechanistic significance and adaptive value to animal cell systems.

Gene order in bacteria is poorly conserved during evolution. For example, although many homologous genes are shared by the proteobacteria Escherichia coli, Haemophilus influenzae and Helicobacter pylori, their relative positions are very different in each genome, except local functional clusters such as operons. The complete sequences of the more closely related bacterial genomes, such as pairs of Chlamydia, H. pylori and Mycobacterium species, now allow identification of the processes and mechanisms involved in genome evolution. Here we provide evidence that a substantial proportion of rearrangements in gene order results from recombination sites that are determined by the positions of the replication forks. Our observations suggest that replication has a major role in directing genome evolution.