Portrait of Dr. William Levison in military uniform Portraits Men

Digital Image

Studio portrait of five Heimann brothers in military uniform during World War I.

l-r: Albert Heimann, Leopold Heimann, Nathan Heimann, Hermann Heimann and Max Heimann; Sons of Julius and Mina Heimann

Mu in vitro transposition applications in protein engineering

Transposons, mobile genetic elements that are ubiquitous in all living organisms have been used as tools in molecular biology for decades. They have the ability to move into discrete DNA locations with no apparent homology to the target site. The utility of transposons as molecular tools is based on their ability to integrate into various DNA sequences efficiently, producing extensive mutant clone libraries that can be used in various molecular biology applications. Bacteriophage Mu is one of the most useful transposons due to its well-characterized and simple in vitro transposition reaction. This study establishes the properties of the Mu in vitro transposition system as a versatile multipurpose tool in molecular biology. In addition, this study describes Mu-based applications for engineering proteins by random insertional transposon mutagenesis in order to study structure-function relationships in proteins. We initially characterized the properties of the minimal Mu in vitro transposition system. We showed that the Mu transposition system works efficiently and accurately and produces insertions into a wide spectrum of target sites in different DNA molecules. Then, we developed a pentapeptide insertion mutagenesis strategy for inserting random five amino acid cassettes into proteins. These protein variants can be used especially for screening important sites for protein-protein interactions. Also, the system may produce temperature-sensitive variants of the protein of interest. Furthermore, we developed an efficient screening system for high-resolution mapping of protein-protein interfaces with the pentapeptide insertion mutagenesis. This was accomplished by combining the mutagenesis with subsequent yeast two-hybrid screening and PCR-based genetic footprinting. This combination allows the analysis of the whole mutant library en masse, without the need for producing or isolating separate mutant clones, and the protein-protein interfaces can be determined at amino acid accuracy. The system was validated by analysing the interacting region of JFC1 with Rab8A, and we show that the interaction is mediated via the JFC1 Slp homology domain. In addition, we developed a procedure for the production of nested sets of N- and C-terminal deletion variants of proteins with the Mu system. These variants are useful in many functional studies of proteins, especially in mapping regions involved in protein-protein interactions. This methodology was validated by analysing the region in yeast Mso1 involved in an interaction with Sec1. The results of this study show that the Mu in vitro transposition system is versatile for various applicational purposes and can efficiently be adapted to random protein engineering applications for functional studies of proteins.

An introduction to ophthalmic biomaterials and their role in tissue engineering and regenerative medicine

This chapter presents a brief history of the development of ophthalmic biomaterials. Particularities in the development of ophthalmic biomaterials are discussed and some of their historic priorities within the general field of biomaterials are revealed or emphasized. The chapter then discusses the role and integration of ophthalmic biomaterials in tissue engineering and regenerative medicine applications.

Fatigue fractures in military conscripts : A study on risk factors, diagnostics and long-term consequences

Fatigue fracture is an overuse injury commonly encountered in military and sports medicine, and known to relate to intensive or recently intensified physical activity. Bone responds to increased stress by enhanced remodeling. If physical stress exceeds bone s capability to remodel, accumulation of microfractures can lead to bone fatigue and stress fracture. Clinical diagnosis of stress fractures is complex and based on patient s anamnesis and radiological imaging. Bone stress fractures are mostly low-risk injuries, healing well after non-operative management, yet, occurring in high-risk areas, stress fractures can progress to displacement, often necessitating surgical treatment and resulting in prolonged morbidity. In the current study, the role of vitamin D as a predisposing factor for fatigue fractures was assessed using serum 25OHD level as the index. The average serum 25OHD concentration was significantly lower in conscripts with fatigue fracture than in controls. Evaluating TRACP-5b bone resorption marker as indicator of fatigue fractures, patients with elevated serum TRACP-5b levels had eight times higher probability of sustaining a stress fracture than controls. Among the 154 patients with exercise induced anterior lower leg pain and no previous findings on plain radiography, MRI revealed a total of 143 bone stress injuries in 86 patients. In 99% of the cases, injuries were in the tibia, 57% in the distal third of the tibial shaft. In patients with injury, forty-nine (57%) patients exhibited bilateral stress injuries. In a 20-year follow-up, the incidence of femoral neck fatigue fractures prior to the Finnish Defence Forces new regimen in 1986 addressing prevention of these fractures was 20.8/100,000, but rose to 53.2/100,000 afterwards, a significant 2.6-fold increase. In nineteen subjects with displaced femoral neck fatigue fractures, ten early local complications (in first postoperative year) were evident, and after the first postoperative year, osteonecrosis of the femoral head in six and osteoarthritis of the hip in thirteen patients were found. It seems likely that low vitamin D levels are related to fatigue fractures, and that an increasing trend exists between TRACP-5b bone resorption marker elevation and fatigue fracture incidence. Though seldom detected by plain radiography, fatigue fractures often underlie unclear lower leg stress-related pain occurring in the distal parts of the tibia. Femoral neck fatigue fractures, when displaced, lead to long-term morbidity in a high percentage of patients, whereas, when non-displaced, they do not predispose patients to subsequent adverse complications. Importantly, an educational intervention can diminish the incidence of fracture displacement by enhancing awareness and providing instructions for earlier diagnosis of fatigue fractures.

Using Water as a Design Element in Crystal Engineering. Host-Guest Compounds of Hydrated 3,5-Dihydroxybenzoic Acid

Thirteen host guest compounds of 3,5-dihydroxybenzoic acid (DHBA) have been structurally characterized. Water molecules occupy the peripheries of a hexagonal void, created with DHBA molecules, and act as ``hooks'' to connect the guest molecules with the host-framework via hydrogen bonding. The ``water hook'' is an OH group acting as a donor. Consequently, the guest molecules were chosen so that they contain good hydrogen bond acceptor functionalities. A number of multicomponent hydrates were isolated with stoichiometries (DHBA)(x)(H2O). (guest),. Of these, compounds with the following as guests were obtained as crystals that were good enough for single crystal work: ethyl acetate (EtOAc), diethyl oxalate, dimethyl oxalate, di(n-propyl) oxalate, diethyl malonate, diethyl succinate, chloroacetonitrile, N,N-dimethyl formamide (DMF), acetone, dimethyl sulfoxide (DMSO), 1-propanol, and 2-butanol. From 2-butanol, a hemihydrate, (DHBA)(2)(H2O), was also obtained concomitantly. Further to guest stabilization, water acts as a good mediator of effective crystal packing and also determines the topology of the host framework. En the present series of compounds, the role of water is wide ranging, and it is not easy to classify it specifically as a host or as a guest.

Studies in crystal engineering. Photochemical and crystallographic investigations of bromocoumarins and (±)-7-(p-bromobenzylidene)piperitone

Studies in crystal engineering. Photochemical and crystallographic investigations of bromocoumarins and (±)-7-(p-bromobenzylidene)piperitone

Role of organic fluorine in crystal engineering

The design of compounds with novel and improved physico-chemical properties as advanced functional materials with a specific application spectrum requires the knowledge about possible supramolecular packing motifs and their experimental control in crystalline lattice. Besides the structure of the individual molecule, non-covalent interactions play a significant role in the determination of molecular conformation, along with the formation of three-dimensional supramolecular architecture in a crystal as a requirement for molecular recognition processes, and the related bioactivity. Involvement of functional groups will contribute to the formation of a predefined packing motif due to their well-defined interactions. The strength and directionality of these interactions create characteristic packing motifs, which can be used for the design of supramolecular arrangements by the development of appropriate strategies for the precise control of their topology. Most relevant of these non-covalent interactions are stacking interactions and hydrogen bonds, which have been subjects of extensive study in the last two decades. In recent literature, substantial efforts have been put in by various researchers towards the understanding of interactions involving organic fluorine and the role they play in generating different packing motifs which guides assembling of molecules in the crystal lattice.

Studies in crystal engineering: structure�reactivity correlations of substituted styrylcoumarins and related systems

The solid state photochemical behaviour of 7-hydroxy-4-styrylcoumarin 1 and several of its derivatives and analogues has been investigated. All the compounds with the exception of 7-methoxy-4-styrylcoumarin 2 are photolabile and yield anti-HT dimers. It has been observed that chloro substitution in the systems studied does not lead to the expected beta-packing mode. The photobehaviour of 1 and 2 has been correlated with their crystal structures. Reasons for alpha-packing have been examined. The systematics in the arrangement of the carbonyl group and phenyl group of the close neighbours in the crystals of 1, 2 and a few other cases are presented.

Reflections on the Hydrogen Bond in Crystal Engineering

Comments on aspects of the new definition of the hydrogen bond specific to crystal engineering are given.

Studies in crystal engineering: Effect of fluorine substitution in crystal packing and topological photodimerization of styryl coumarins in the solid state

Styryl coumarins generally yield centrosymmetric (alpha-mode, anti-HT) photodimers when subjected to irradiation in the solid state, However, the substitution of fluorine dramatically alters the packing mode and steers the molecules 4-(4-fluorostyryl)coumarin 1 and 4-(2-fluorostyryl)coumarin 2 to form a stereospecific photodimer, beta-mode, syn-HH across the styrenic double bond (yield 78-85%). The stereochemistry of the photodimer 2a has been established by X-ray crystallography. There is no evidence for the presence of C-H ... F interactions. The true nature of the weak atom-atom interactions called into play when fluorine is substituted is not clear, It is observed that the fluoro substituted compounds have greater crystal density than the corresponding unsubstituted ones.

Studies in crystal engineering: Crystal packing, topological photodimerization and structure-reactivity correlations in fluoro-substituted styrylcoumarins

In continuation of our studies on the influence of fluoro substitution on the solid state photobehaviour and packing pattern of styrylcoumarins, the results obtained for 4-(3-fluorostyryl)coumarin 1, 4-styryl-6-fluorocoumarin 2 and 4-styryl-7-fluorocoumarin 3 are presented. The configuration of the dimers was established on the basis of crystal packing of 1 and 2 (alpha-packed). A rationale for the significantly lower dimer yield in the crystal for 2 is proposed. In the observed centrosymmetric arrangement of the reactants the C=O ...pi (phenyl) contacts seem to provide additional attractive interactions. C-H ... O and C-H ... F hydrogen bonding seems to provide stability in these structures.

Studies in crystal engineering: Steering ability of fluorine in styrylcoumarins

In continuation of our studies on crystal engineering using fluorine as a steering group, the photobehaviour of di and tri fluoro 4-styrylcoumarins has been examined. It is found that out of the five derivatives, four crystallize into P-packing mode producing syn-HH photodimer upon irradiation whereas the parent hydrocarbon produces an anti K-T dimer. The packing features of the photolabile crystals of 4-(4-fluorostyryl)-6-fluorocoumarin (1), 4-(2,6-difluorostyryl) 6-fluorocoumarin (2) and the photodimer (3a) of 4-(2,6-fluorostyryl)-7-fluorocoumarin (3) have been determined by single crystal X-ray diffraction studies. The stereochemistry of the photodimer of 4-(2-fluorostyryl)-6-fluorocoumarin (4) is deduced based on preliminary X-ray crystallographic data. However, 4-(2,6-difluorostyryl) coumarin (5) is photoinert. The remarkable steering ability of fluorine is established with the molecular packing in the crystal lattice leading to the formation of syn H-H dimer in the above four examples. (C) 1999 Elsevier Science Ltd. All rights reserved.