In contrast to replicative DNA polymerases, Sulfolobus solfataricus Dpo4 showed a limited decrease in catalytic efficiency (k cat /K m ) for insertion of dCTP opposite a series of N 2 -alkylguanine templates of increasing size from (methyl (Me) to (9-anthracenyl)-Me (Anth)). Fidelity was maintained with increasing size up to (2-naphthyl)-Me (Naph). The catalytic efficiency increased slightly going from the N 2 -NaphG to the N 2 -AnthG substrate, at the cost of fidelity. Pre-steady-state kinetic bursts were observed for dCTP incorporation throughout the series (N 2 -MeG to N 2 -AnthG), with a decrease in the burst amplitude and k pol , the rate of single-turnover incorporation. The presteady-state kinetic courses with G and all of the six N 2 -alkyl G adducts could be fit to a general DNA polymerase scheme to which was added an inactive complex in equilibrium with the active ternary Dpo4⅐DNA⅐dNTP complex, and only the rates of equilibrium with the inactive complex and phosphodiester bond formation were altered. Two crystal structures of Dpo4 with a template N 2 -NaphG (in a post-insertion register opposite a 3-terminal C in the primer) were solved. One showed N 2 -NaphG in a syn conformation, with the naphthyl group located between the template and the Dpo4 "little finger" domain. The Hoogsteen face was within hydrogen bonding distance of the N4 atoms of the cytosine opposite N 2 -NaphG and the cytosine at the ؊2 position. The second structure showed N 2 -Naph G in an anti conformation with the primer terminus largely disordered. Collectively these results explain the versatility of Dpo4 in bypassing bulky G lesions.

The container relocation problem, where containers that are stored in bays are retrieved in a fixed sequence, is a crucial port operation. Existing approaches using branch and bound algorithms are only able to optimally solve small cases in a practical time frame. In this paper, we investigate iterative deepening A* algorithms (rather than branch and bound) using new lower bound measures and heuristics, and show that this approach is able to solve much larger instances of the problem in a time frame that is suitable for practical application. We also examine a more difficult variant of the problem that has been largely ignored in existing literature.Note to Practitioners-Container retrieval is an important operation in a container port. When a ship arrives, containers stored in the port yard are first retrieved by yard crane, loaded onto autoguided vehicles, transported to quay cranes, and loaded onto the ship by quay crane. Due to various operational constraints, e.g., maintenance of vessel balance and safety issues, the containers in a storage bay are retrieved one by one in a fixed sequence. When the next container to be retrieved is not at the top of its stack, all other containers above it must then be first relocated onto other stacks within the bay. The relocation of a container is a time-consuming operation that essentially dominates all other aspects of the problem, and therefore it is important that the retrieval plan minimizes the number of such relocations. This study proposes a method to generate a near-optimal retrieval plan for yard cranes. This often arises as a subproblem when devising an overall plan for port operations that maximizes throughput, which involves the coordination of multiple pieces of machinery. Our approach produces significantly better results than all existing approaches.Index Terms-Container relocation problem, container yard operation, iterative deepening A*.

This article reviews the recent progress made in asymmetric catalysis in the nanopores of mesoporous materials and periodic mesoporous organosilicas (PMOs). Some examples of chiral catalysts within the nanopores show improved catalytic performance compared to homogeneous catalysts. The factors including the confinement effect, the properties of the linkages and the microenvironment in nanopores, which affect the activity and enantioselectivity of asymmetric catalysis in nanopores, are discussed.

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Interactions between gene 4 helicase and gene 5 DNA polymerase (gp5) are crucial for leading-strand DNA synthesis mediated by the replisome of bacteriophage T7. Interactions between the two proteins that assure high processivity are known but the interactions essential to initiate the leading-strand DNA synthesis remain unidentified. Replacement of solution-exposed basic residues (K587, K589, R590, and R591) located on the front surface of gp5 with neutral asparagines abolishes the ability of gp5 and the helicase to mediate strand-displacement synthesis. This front basic patch in gp5 contributes to physical interactions with the acidic C-terminal tail of the helicase. Nonetheless, the altered polymerase is able to replace gp5 and continue ongoing strand-displacement synthesis. The results suggest that the interaction between the C-terminal tail of the helicase and the basic patch of gp5 is critical for initiation of strand-displacement synthesis. Multiple interactions of T7 DNA polymerase and helicase coordinate replisome movement.DNA polymerase-helicase interaction | strand-displacement DNA synthesis | T7 bacteriophage | T7 replisome B acteriophage T7 has a simple and efficient DNA replication system whose basic reactions mimic those of more complex replication systems (1). The T7 replisome consists of gene 5 DNA polymerase (gp5), the processivity factor, Escherichia coli thioredoxin (trx), gene 4 helicase-primase (gp4), and gene 2.5 ssDNA binding protein (gp2.5) (Fig. 1A). Gp5 forms a high-affinity complex with trx (gp5/trx) to increase the processivity of nucleotide polymerization (2). The C-terminal helicase domain of gp4 assembles as a hexamer and unwinds dsDNA to produce two ssDNA templates for leading-and lagging-strand gp5/trx. The N-terminal primase domain of gp4 catalyzes the synthesis of tetraribonucleotides that are used as primers for the lagging-strand gp5/trx. This gp5/trx also binds to helicase to form a replication loop containing the nascent Okazaki fragment. Gp2.5 coats the ssDNA to remove secondary structures and it also physically interacts with gp4 and gp5/trx, interactions essential for coordination of leading and lagging-strand synthesis (3).Other DNA replication systems are generally more complicated than the T7 system. In E. coli, at least 13 proteins are required for a functional replisome and eight proteins are required in bacteriophage T4 infected cells (1, 4, 5). The two essential helicase and primase are separate proteins although they must physically interact to properly function (5). The additional proteins include processivity clamps and loading proteins. The existing T7 gp4 has usurped helicase and primase functions. The proofreading exonuclease activity resides within the N-terminal portion of gp5, whereas in the E. coli system it resides within ϵ subunit of the polymerase holoenzyme (5). Trx not only binds to gp5 to increase processivity but it also configures the trx-binding loop in gp5 for the binding of gp2.5 and gp4 (6). An interaction of the C-terminal tail of gp2.5 with...

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