Structural analysis of the activation-induced deoxycytidine deaminase required in immunoglobulin diversification.

Activation-induced deoxycytidine deaminase (AID) initiates somatic hypermutation (SHM) and class-switch recombination (CSR) by deaminating C→U during transcription of Ig-variable (V) and Ig-switch (S) region DNA, which is essential to produce high-affinity antibodies. Here we report the crystal structure of a soluble human AID variant at 2.8Å resolution that favors targeting WRC motifs (W=A/T, R=A/G) in vitro, and executes Ig V SHM in Ramos B-cells.

Random-walk enzymes.

Enzymes that rely on random walk to search for substrate targets in a heterogeneously dispersed medium can leave behind complex spatial profiles of their catalyzed conversions. The catalytic signatures of these random-walk enzymes are the result of two coupled stochastic processes: scanning and catalysis. Here we develop analytical models to understand the conversion profiles produced by these enzymes, comparing an intrusive model, in which scanning and catalysis are tightly coupled, against a loosely coupled passive model.

A mathematical model for scanning and catalysis on single-stranded DNA, illustrated with activation-induced deoxycytidine deaminase.

We formulated a master equation-based mathematical model to analyze random scanning and catalysis for enzymes that act on single-stranded DNA (ssDNA) substrates. Catalytic efficiencies and intrinsic scanning distances are deduced from the distribution of positions and gap lengths between a series of catalytic events occurring over time, which are detected as point mutations in a lacZα-based reporter sequence containing enzyme target motifs. Mathematical analysis of the model shows how scanning motions become separable from the catalysis when the proper statistical properties of the mutation pattern are used to interpret the readouts.

Mutational analysis of an archaeal minichromosome maintenance protein exterior hairpin reveals critical residues for helicase activity and DNA binding.

Background: The mini-chromosome maintenance protein (MCM) complex is an essential replicative helicase for DNA replication in Archaea and Eukaryotes. While the eukaryotic complex consists of six homologous proteins (MCM2-7), the archaeon Sulfolobus solfataricus has only one MCM protein (ssoMCM), six subunits of which form a homohexamer. We have recently reported a 4.