A proposed concept for a crustal dynamics information management network

The findings of a requirements and feasibility analysis of the present and potential producers, users, and repositories of space-derived geodetic information are summarized. A proposed concept is presented for a crustal dynamics information management network that would apply state of the art concepts of information management technology to meet the expanding needs of the producers, users, and archivists of this geodetic information

\u3cem\u3ePlasmodium falciparum\u3c/em\u3e SSB Tetramer Binds Single-Stranded DNA Only in a Fully Wrapped Mode

The tetrameric Escherichia coli single-stranded DNA (ssDNA) binding protein (Ec-SSB) functions in DNA metabolism by binding to ssDNA and interacting directly with numerous DNA repair and replication proteins. Ec-SSB tetramers can bind ssDNA in multiple DNA binding modes that differ in the extent of ssDNA wrapping. Here, we show that the structurally similar SSB protein from the malarial parasite Plasmodium falciparum (Pf-SSB) also binds tightly to ssDNA but does not display the same number of ssDNA binding modes as Ec-SSB, binding ssDNA exclusively in fully wrapped complexes with site sizes of 52–65 nt/tetramer. Pf-SSB does not transition to the more cooperative (SSB)35 DNA binding mode observed for Ec-SSB. Consistent with this, Pf-SSB tetramers also do not display the dramatic intra-tetramer negative cooperativity for binding of a second (dT)35 molecule that is evident in Ec-SSB. These findings highlight variations in the DNA binding properties of these two highly conserved homotetrameric SSB proteins, and these differences might be tailored to suit their specific functions in the cell

Generic functional requirements for a NASA general-purpose data base management system

Generic functional requirements for a general-purpose, multi-mission data base management system (DBMS) for application to remotely sensed scientific data bases are detailed. The motivation for utilizing DBMS technology in this environment is explained. The major requirements include: (1) a DBMS for scientific observational data; (2) a multi-mission capability; (3) user-friendly; (4) extensive and integrated information about data; (5) robust languages for defining data structures and formats; (6) scientific data types and structures; (7) flexible physical access mechanisms; (8) ways of representing spatial relationships; (9) a high level nonprocedural interactive query and data manipulation language; (10) data base maintenance utilities; (11) high rate input/output and large data volume storage; and adaptability to a distributed data base and/or data base machine configuration. Detailed functions are specified in a top-down hierarchic fashion. Implementation, performance, and support requirements are also given

A Closed Network Queue Model of Underground Coal Mining Production, Failure, and Repair

Underground coal mining system production, failures, and repair cycles were mathematically modeled as a closed network of two queues in series. The model was designed to better understand the technological constraints on availability of current underground mining systems, and to develop guidelines for estimating the availability of advanced mining systems and their associated needs for spares as well as production and maintenance personnel. It was found that: mine performance is theoretically limited by the maintainability ratio, significant gains in availability appear possible by means of small improvements in the time between failures the number of crews and sections should be properly balanced for any given maintainability ratio, and main haulage systems closest to the mine mouth require the most attention to reliability

Multiple C-Terminal Tails within a Single \u3cem\u3eE. coli\u3c/em\u3e SSB Homotetramer Coordinate DNA Replication and Repair

Escherichia coli single-stranded DNA binding protein (SSB) plays essential roles in DNA replication, recombination and repair. SSB functions as a homotetramer with each subunit possessing a DNA binding domain (OB-fold) and an intrinsically disordered C-terminus, of which the last nine amino acids provide the site for interaction with at least a dozen other proteins that function in DNA metabolism. To examine how many C-termini are needed for SSB function, we engineered covalently linked forms of SSB that possess only one or two C-termini within a four-OB-fold “tetramer”. Whereas E. coli expressing SSB with only two tails can survive, expression of a single-tailed SSB is dominant lethal. E. coli expressing only the two-tailed SSB recovers faster from exposure to DNA damaging agents but accumulates more mutations. A single-tailed SSB shows defects in coupled leading and lagging strand DNA replication and does not support replication restart in vitro. These deficiencies in vitro provide a plausible explanation for the lethality observed in vivo. These results indicate that a single SSB tetramer must interact simultaneously with multiple protein partners during some essential roles in genome maintenance

Intrinsically Disordered C-Terminal Tails of \u3cem\u3eE. coli\u3c/em\u3e Single-Stranded DNA Binding Protein Regulate Cooperative Binding to Single-Stranded DNA

The homotetrameric Escherichia coli single-stranded DNA binding protein (SSB) plays a central role in DNA replication, repair and recombination. E. coli SSB can bind to long single-stranded DNA (ssDNA) in multiple binding modes using all four subunits [(SSB)65 mode] or only two subunits [(SSB)35 binding mode], with the binding mode preference regulated by salt concentration and SSB binding density. These binding modes display very different ssDNA binding properties with the (SSB)35 mode displaying highly cooperative binding to ssDNA. SSB tetramers also bind an array of partner proteins, recruiting them to their sites of action. This is achieved through interactions with the last 9 amino acids (acidic tip) of the intrinsically disordered linkers (IDLs) within the four C-terminal tails connected to the ssDNA binding domains. Here, we show that the amino acid composition and length of the IDL affects the ssDNA binding mode preferences of SSB protein. Surprisingly, the number of IDLs and the lengths of individual IDLs together with the acidic tip contribute to highly cooperative binding in the (SSB)35 binding mode. Hydrodynamic studies and atomistic simulations suggest that the E. coli SSB IDLs show a preference for forming an ensemble of globular conformations, whereas the IDL from Plasmodium falciparum SSB forms an ensemble of more extended random coils. The more globular conformations correlate with cooperative binding

Velocity and processivity of helicase unwinding of double-stranded nucleic acids

Helicases are molecular motors which unwind double-stranded nucleic acids (dsNA) in cells. Many helicases move with directional bias on single-stranded (ss) nucleic acids, and couple their directional translocation to strand separation. A model of the coupling between translocation and unwinding uses an interaction potential to represent passive and active helicase mechanisms. A passive helicase must wait for thermal fluctuations to open dsNA base pairs before it can advance and inhibit NA closing. An active helicase directly destabilizes dsNA base pairs, accelerating the opening rate. Here we extend this model to include helicase unbinding from the nucleic-acid strand. The helicase processivity depends on the form of the interaction potential. A passive helicase has a mean attachment time which does not change between ss translocation and ds unwinding, while an active helicase in general shows a decrease in attachment time during unwinding relative to ss translocation. In addition, we describe how helicase unwinding velocity and processivity vary if the base-pair binding free energy is changed.Comment: To appear in special issue on molecular motors, Journal of Physics - Condensed Matte

Is a fully wrapped SSB-DNA complex essential for Escherichia coli survival?

Escherichia coli single-stranded DNA binding protein (SSB) is an essential homotetramer that binds ssDNA and recruits multiple proteins to their sites of action during genomic maintenance. Each SSB subunit contains an N-terminal globular oligonucleotide/oligosaccharide binding fold (OB-fold) and an intrinsically disordered C-terminal domain. SSB binds ssDNA in multiple modes in vitro, including the fully wrapped (SSB)(65) and (SSB)(56) modes, in which ssDNA contacts all four OB-folds, and the highly cooperative (SSB)(35) mode, in which ssDNA contacts an average of only two OB-folds. These modes can both be populated under physiological conditions. While these different modes might be used for different functions, this has been difficult to assess. Here we used a dimeric SSB construct with two covalently linked OB-folds to disable ssDNA binding in two of the four OB-folds thus preventing formation of fully wrapped DNA complexes in vitro, although they retain a wild-type-like, salt-dependent shift in cooperative binding to ssDNA. These variants complement wild-type SSB in vivo indicating that a fully wrapped mode is not essential for function. These results do not preclude a normal function for a fully wrapped mode, but do indicate that E. coli tolerates some flexibility with regards to its SSB binding modes

Allosteric effects of E. coli SSB and RecR proteins on RecO protein binding to DNA

Escherichia coli single stranded (ss) DNA binding protein (SSB) plays essential roles in DNA maintenance. It binds ssDNA with high affinity through its N-terminal DNA binding core and recruits at least 17 different SSB interacting proteins (SIPs) that are involved in DNA replication, recombination, and repair via its nine amino acid acidic tip (SSB-Ct). E. coli RecO, a SIP, is an essential recombination mediator protein in the RecF pathway of DNA repair that binds ssDNA and forms a complex with E. coli RecR protein. Here, we report ssDNA binding studies of RecO and the effects of a 15 amino acid peptide containing the SSB-Ct monitored by light scattering, confocal microscope imaging, and analytical ultracentrifugation (AUC). We find that one RecO monomer can bind the oligodeoxythymidylate, (dT)15, while two RecO monomers can bind (dT)35 in the presence of the SSB-Ct peptide. When RecO is in molar excess over ssDNA, large RecO-ssDNA aggregates occur that form with higher propensity on ssDNA of increasing length. Binding of RecO to the SSB-Ct peptide inhibits RecO-ssDNA aggregation. RecOR complexes can bind ssDNA via RecO, but aggregation is suppressed even in the absence of the SSB-Ct peptide, demonstrating an allosteric effect of RecR on RecO binding to ssDNA. Under conditions where RecO binds ssDNA but does not form aggregates, SSB-Ct binding enhances the affinity of RecO for ssDNA. For RecOR complexes bound to ssDNA, we also observe a shift in RecOR complex equilibrium towards a RecR4O complex upon binding SSB-Ct. These results suggest a mechanism by which SSB recruits RecOR to facilitate loading of RecA onto ssDNA gaps