Structure of the tetrameric restriction endonuclease NgoMIV in complex with cleaved DNA.

The crystal structure of the NgoMIV restriction endonuclease in complex with cleaved DNA has been determined at 1.6 A resolution. The crystallographic asymmetric unit contains a protein tetramer and two DNA molecules cleaved at their recognition sites. This is the first structure of a tetrameric restriction enzyme-DNA complex. In the tetramer, two primary dimers are arranged back to back with two oligonucleotides bound in clefts on opposite sides of the tetramer. The DNA molecules retain a B-type conformation and have an enclosed angle between their helical axes of 60 degrees. Sequence-specific interactions occur in both the major and minor grooves. Two Mg2+ ions are located close to the cleaved phosphate at the active site of NgoMIV. Biochemical experiments show that interactions between the recognition sites within the tetramer greatly increase DNA cleavage efficiency.

Crystal structure of MunI restriction endonuclease in complex with cognate DNA at 1.7 A resolution.

The MunI restriction enzyme recognizes the palindromic hexanucleotide sequence C/AATTG (the '/' indicates the cleavage site). The crystal structure of its active site mutant D83A bound to cognate DNA has been determined at 1.7 A resolution. Base-specific contacts between MunI and DNA occur exclusively in the major groove. While DNA-binding sites of most other restriction enzymes are comprised of discontinuous sequence segments, MunI combines all residues involved in the base-specific contacts within one short stretch (residues R115-R121) located at the N-terminal region of the 3(10)4 helix. The outer CG base pair of the recognition sequence is recognized solely by R115 through hydrogen bonds made by backbone and side chain atoms to both bases. The mechanism of recognition of the central AATT nucleotides by MunI is similar to that of EcoRI, which recognizes the G/AATTC sequence. The local conformation of AATT deviates from the typical B-DNA form and is remarkably similar to EcoRI-DNA. It appears to be essential for specific hydrogen bonding and recognition by MunI and EcoRI.

Skiing injuries in children, adolescents, and adults.

We prospectively gathered data on skiing injuries that had been sustained at the Sugarbush North ski area since 1972 and at the Sugarbush South ski area since 1981. The purpose of the current study was to document the overall rates of injury in children, adolescents, and adults participating in alpine skiing. We also sought to determine the ten most common injuries in each age-group. Finally, we analyzed short-term and long-term trends to determine if changes in equipment had had an effect on the frequency or pattern of injury. From the 1981-1982 to the 1993-1994 season, there were 2.79 injuries per 1000 skier days: 4.27 injuries in children, 2.93 in adolescents, and 2.69 in adults. During the last eight years of the study, the most common injuries were a contusion of the knee in children, a sprain of the ulnar collateral ligament of the thumb in adolescents, and a grade-III sprain of the anterior cruciate ligament in adults. The short-term trends revealed that, in children, the frequency of tibial fractures decreased 10 per cent while that of fractures of the upper extremity increased 8 per cent. The long-term trends showed that, in adults, the rate of tibial fractures decreased 89 per cent while that of injuries of the anterior cruciate ligament increased 280 per cent. The overall rate of injury decreased 43 per cent from the beginning of the study in 1972 to the end of the study in 1994; the decrease was 58 per cent in children, 45 per cent in adolescents, and 42 per cent in adults. Data on the types of equipment and the binding-release values were collected prospectively from injured skiers and from 2083 non-injured skiers. Of the fifty-nine skiers who sustained a spiral fracture of the tibia, forty-two (71 per cent) had binding-release values that were higher than the average for the uninjured group. We believe that the use of properly functioning modern equipment will decrease the rate of injury, particularly in children.

Ion resonance electromagnetic field stimulation of fracture healing in rabbits with a fibular ostectomy.

Rabbits with a fibular ostectomy were exposed for 28 days to magnetic fields that satisfied the ion resonance conditions for calcium or magnesium. The rabbits were exposed to whole body treatment for 1/2 hour, 3 hours, or 24 hours per day. The fibulae from the experimental and control animals were removed surgically and were subjected to force-deflection testing to establish the stiffness of the healed fracture. The fibulae from the rabbits exposed to the ion resonance magnetic fields were found to be 55-299% (p < 0.01) more robust than the fibulae from the control animals.