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Biomedical subjects

J H Shim

Publications and source records attributed to J H Shim.

At least 19 recordsLinked to original sources
PubMed2000

Design, synthesis, and biological evaluation of fluoronitrophenyl substituted folate analogues as potential inhibitors of GAR transformylase and AICAR transformylase.

The examination results of a novel series of potential inhibitors of glycinamide ribonucleotide transformylase (GAR Tfase) and aminoimidazole carboxamide transformylase (AICAR Tfase) are reported. These agents incorporate an electrophilic fluoronitrophenyl group that can potentially react with an active site nucleophile or the substrate GAR/AICAR amine via nucleophilic aromatic substitution.

Animals↗
PubMed1999

Catalytic mechanism of Escherichia coli glycinamide ribonucleotide transformylase probed by site-directed mutagenesis and pH-dependent studies.

Site-directed mutagenesis followed by studies of the pH dependence of the kinetic parameters of the mutants has been used to probe the role of the active site residues and loops in catalysis by glycinamide ribonucleotide transformylase (EC 2.1.2.2). The analysis of the mutants of the strictly conserved active site residues, His108 and Asp144, revealed that His108 acts in a salt bridge with Asp144 as a general acid catalyst with a pK(a) value of 9.7. Asp144 also plays a key role in the preparation of the active site geometry for catalysis. The rate-limiting step in the pH range of 6-10 appears to be the catalytic steps involving tetrahedral intermediates, supported by the observation of a pL (L being H or D)-independent solvent deuterium isotope effect of 2. The ionization of the amino group of glycinamide ribonucleotide both as a free and as a bound form dominates the kinetic behavior at low pH. The analysis of a mutation, H121Q, within the loop spanning amino acids 111-131 suggests the closure of the loop is involved in the binding of the substrate. The kinetic behavior parallels pH effects revealed by a series of X-ray crystallographic structures of the apoenzyme and inhibitor-bound enzyme [Su, Y., Yamashita, M. M., Greasley, S. E. , Mullen, C. A., Shim, J. H., Jennings, P. A., Benkovic, S. J., and Wilson, I. A. (1998) J. Mol. Biol. 281, 485-499], permitting a more exact formulation of the probable catalytic mechanism.

Binding Sites↗
PubMed1999

Combinatorial protein engineering by incremental truncation.

We have developed a combinatorial approach, using incremental truncation libraries of overlapping N- and C-terminal gene fragments, that examines all possible bisection points within a given region of an enzyme that will allow the conversion of a monomeric enzyme into its functional heterodimer. This general method for enzyme bisection will have broad applications in the engineering of new catalytic functions through domain swapping and chemical synthesis of modified peptide fragments and in the study of enzyme evolution and protein folding. We have tested this methodology on Escherichia coli glycinamide ribonucleotide formyltransferase (PurN) and, by genetic selection, identified PurN heterodimers capable of glycinamide ribonucleotide transformylation. Two were chosen for physical characterization and were found to be comparable to the wild-type PurN monomer in terms of stability to denaturation, activity, and binding of substrate and cofactor. Sequence analysis of 18 randomly chosen, active PurN heterodimers revealed that the breakpoints primarily clustered in loops near the surface of the enzyme, that the breaks could result in the deletion of highly conserved residues and, most surprisingly, that the active site could be bisected.

Amino Acid Sequence↗
PubMed1999

A combinatorial approach to hybrid enzymes independent of DNA homology.

We present a methodology, termed incremental truncation for the creation of hybrid enzymes (ITCHY), that creates combinatorial fusion libraries between genes in a manner that is independent of DNA homology. We compared the ability of ITCHY and DNA shuffling to create interspecies fusion libraries between fragments of the Escherichia coli and human glycinamide ribonucleotide transformylase genes, which have only 50% identity on the DNA level. Sequencing of several randomly selected positives from each library illustrated that ITCHY identified a more diverse set of active fusion points including those in regions of nonhomology and those with crossover points that diverged from the sequence alignment. Furthermore, some of the hybrids found by ITCHY that were fused at nonhomologous locations had activities that were greater than or equal to the activity of the hybrids found by DNA shuffling.

Amino Acid Sequence↗
PubMed1999

Muscle activation during the tennis volley.

PURPOSE: To broaden our understanding of muscle function during the tennis volley under different ball placement and speed conditions by examining the activity of selected superficial muscles of the stroking arm and shoulder (flexor carpi radialis, extensor carpi radialis, triceps brachii, deltoids, and pectoralis major) and muscles related to postural support (left and right external oblique, lumbar erector spinae, and gastrocnemius) during the volley. METHODS: Seven skilled tennis players were asked to perform volley strokes under 18 experimental conditions, including variations in lateral contact location (forehand and backhand), ball contact height (high, middle, and low), and ball speed (fast, medium, and slow). A ball machine was modified so that the subjects could not predict the ball trajectory before it was released from the machine. Muscle activity was determined using surface electromyographic (EMG) techniques, and the critical instants of a volley were determined using two force platforms and two high-speed (120 Hz) video cameras. Average EMG values for different phases of the volley, defined by the critical instants, were computed. RESULTS AND CONCLUSIONS: In general, muscle activity increased with increasing ball speed. The extensor carpi radialis was more active than the flexor carpi radialis during both forehand and backhand volleys, suggesting the importance of wrist extension/abduction and grip strength. The increase in EMG levels in the forearm muscles shortly before the ball impact indicated that the subjects did not tighten their grip and wrist until moments before ball impact. Both antero-middle and postero-middle deltoids were active in most stroke phases. However, the roles of the deltoid muscles during a volley cannot be determined without knowing the actions of the other shoulder joint muscles.

Adolescent↗
PubMed1999

Movement characteristics of the tennis volley.

PURPOSE: The purpose of this study was to examine selected movement characteristics of the tennis volley by evaluating temporal and ground reaction force (GRF) parameters. METHODS: Seven skilled tennis players performed volley strokes under 18 experimental conditions including variations in lateral contact location (forehand (FH) and backhand (BH)), ball contact height (high, middle, low), and ball speed (fast, medium, slow). A ball machine was modified so that the subjects could not predict the ball trajectory before it was released from the machine. The GRF and temporal parameters were determined using two force platforms and two high-speed (120 Hz) video cameras, respectively. Average and maximum values of each measured parameter were computed for different phases of the volley. RESULTS: The average reaction times (from ball release to initial racquet movement (IRM)) for FH and BH trials were 226 and 205 ms, respectively, and the difference was statistically significant. The average stroke time (from IRM to ball impact) ranged from 381 ms in fast speed trials to 803 ms in slow speed trials. A distinct racquet forward motion immediately before ball impact occurred in 75% of the trials and they were evenly distributed between FH and BH trials. An ipsilateral side step (a side step of the foot on the same side of the oncoming ball before the crossover step of the other foot) occurred more often in FH (45%) than in BH (34%) trials. CONCLUSIONS: The GRF during the stroke phase suggest that the subjects initiated lateral movement by leaning sideward when ball velocity was low and by a vigorous pushoff of the contralateral foot when ball velocity was high.

Adolescent↗
PubMed1999

Limited stress response in Streptococcus pneumoniae.

In Streptococcus pneumoniae, heat shock induces the synthesis of 65-, 73-, and 84-kDa proteins, and ethanol shock induces a 104-kDa protein. In this study, the 65-, 84-, and 104-kDa proteins were identified as members of the GroEL, ClpL and alcohol dehydrogenase families, respectively, and the general properties of the stress response of S. pneumoniae to several other stresses were characterized. However, several stresses which are known to induce stress responses in Escherichia coli and Bacillus subtilis failed to induce any high molecular weight heat-shock proteins (HSPs) such as GroEL and DnaK homologues. A minor temperature shift from 30 to 37 C triggered induction of the homologues of DnaK and GroEL of E. coli. These features may provide a foundation for evaluating the role of heat-shock proteins relative to the physiology and pathogenesis of pneumococcus.

Amino Acid Sequence↗
PubMed1998

A pH-dependent stabilization of an active site loop observed from low and high pH crystal structures of mutant monomeric glycinamide ribonucleotide transformylase at 1.8 to 1.9 A.

A mutation in the dimer interface of Escherichia coli glycinamide ribonucleotide transformylase (GarTfase) disrupts the observed pH-dependent association of the wild-type enzyme, but has no observable effect on the enzyme activity. Here, we assess whether a pH effect on the enzyme's conformation is sufficient by itself to explain the pH-dependence of the GarTfase reaction. A pH-dependent conformational change is observed between two high-resolution crystal structures of the Glu70Ala mutant GarTfase at pH 3.5 (1.8 A) and 7.5 (1.9 A). Residues 110 to 131 in GarTfase undergo a transformation from a disordered loop at pH 3.5, where the enzyme is inactive, to an ordered loop-helix structure at pH 7.5, where the enzyme is active. The ordering of this flexible loop-helix has a direct effect on catalytic residues in the active site, binding of the folate cofactor and shielding of the active site from solvent. A main-chain carbonyl oxygen atom from Tyr115 in the ordered loop forms a hydrogen bond with His108, and thereby provides electronic and structural stabilization of this key active site residue. Kinetic data indicate that the pKa of His108 is in fact raised to 9. 2. The loop movement can be correlated with elevation of the His pKa, but with further stabilization, probably from Asp144, after the binding of folate cofactor. Leu118, also in the loop, becomes positioned near the p-amino benzoic acid binding site, providing additional hydrophobic interactions with the cofactor 10-formyl tetrahydrofolate. Thus, the pH-dependence of the enzyme activity appears to arise from local active site rearrangements and not from differences due to monomer-dimer association.

Binding Sites↗
PubMed1998

Evaluation of the kinetic mechanism of Escherichia coli glycinamide ribonucleotide transformylase.

A kinetic scheme is presented for Escherichia coli glycinamide ribonucleotide transformylase (GAR transformylase, EC 2.1.2.2) based on a steady-state and pre-steady-state kinetic analysis of the reaction in both directions employing stopped-flow absorbance and fluorescence spectroscopy. Steady-state parameters showed that kcat for the reverse direction is about 10 times lower than that for the forward direction although the Km values for formyl dideazafolate and dideazafolate or for glycinamide ribonucleotide and formyl glycinamide ribonucleotide are similar. No pre-steady-state transient was observed in either direction, and the single-turnover rate constant under saturating levels of substrates in each direction was found to be very close to the respective steady-state kcat value. This indicates that steps involving ternary complexes are rate-determining for steady-state turnover in each direction. By conducting the single-turnover reactions under various preincubation and mixing conditions, a random sequential kinetic mechanism was implicated in which the enzyme binds glycinamide ribonucleotide or formyl dideazafolate productively in no obligatory order. The collective data provided a quantitative kinetic scheme to serve as a basis for the analysis of mutations.

Bacterial Proteins↗
PubMed1998

Accumulation of tributyl- and triphenyltin compounds in Pacific oyster, Crassostrea gigas, from the Chinhae Bay System, Korea.

Butyl- and phenyltin residues were quantified in seawater and Pacific oyster (Crassostrea gigas) from the Chinhae Bay System, Korea. Butyltin compounds were detected in all the seawater and C. gigas samples, whereas phenyltin compounds were not detected in any seawater samples. Tributyltin (TBT) concentrations in seawater ranged <8-35 ng Sn L-1. TBT and triphenyltin (TPhT) concentrations in oysters ranged 95-885 and 155-678 ng Sn g-1, respectively. Spatial distribution of TBT was closely related to boating and dry-docking activities. However, spatial distribution of TPhT was not consistent with that of TBT. The estimated biological concentration factor (BCF) for TBT in C. gigas was about 25,000. Furthermore, 19 and 28% of total body burdens of TBT and TPhT were found in gonadal mass of C. gigas just prior to the spawning period, indicating that a proportional amount of TBT and TPhT would be released with a following reproductive process.

Animals↗
PubMed1998

Wrist-driven prehension prosthesis for amputee patients with disarticulation of the thumb and index finger.

Partial hand amputations may leave significant functional limitations that are difficult to ameliorate by either orthoses or prostheses. Disarticulation of the thumb and index finger at the metacarpophalangeal joint level and the range of motion of the three remaining fingers would be insufficient to provide any type of grasp. In cases like this, a cosmetic hand usually seems to be the best solution. This report describes the design and use of a wrist-driven prehension prosthesis that was applied to a patient and provided a considerable improvement in function and cosmesis.

Amputation, Traumatic↗
PubMed1997

Histopathological findings, viral DNA distribution and lymphocytic immunophenotypes in vesicular and papular types of herpes zoster.

The characteristics rash of herpes zoster begins as erythematous macules and papules, progressing to vesicles within 12-24 h. Patients with persistent papules without vesicular change are occasionally found. Our aim was to elucidate differences in vesicular and papular types of herpes zoster. Biopsy specimens from 21 patients were examined by an in situ hybridization method to observe viral distribution, and lymphocytic immunophenotypes were evaluated immunohistochemically. There was no differences in cell-mediated immunity or immunophenotypes in lymphocytic infiltrates between vesicular and papular types of herpes zoster. DNA of varicella-zoster virus was detected in the epidermis and hair follicles in the vesicular type but was found in the pilosebaceous unit in the papular type. This indicates that the appearance of clinical types of herpes zoster depends on the infected site of varicella-zoster virus in the tissue.

Adolescent↗
PubMed1996

A case of Cobb syndrome associated with lymphangioma circumscriptum.

Cobb syndrome describes the association of a spinal angioma and a cutaneous angioma such as nevus flammeus or angiokeratoma of the corresponding dermatome. A 23-year-old Korean female showed spiral cord angiomas and segmentally distributed nevus flammeus of corresponding dermatomes (Th5-Th2). Interestingly she had a lymphangioma circumscriptum in the same dermatomes. To the best of our knowledge, there is no report of this constellation. Therefore, we suggest that lymphangioma circumscriptum may be considered as a skin finding of Cobb syndrome.

Adult↗
PubMed1995

Trichodiene synthase. Substrate specificity and inhibition.

The substrate specificity of the sesquiterpene synthase trichodiene synthase was examined by determining the Vmax and Km parameters for the natural substrate, trans,trans-farnesyl diphosphate (1), its stereoisomer, cis,trans-farnesyl diphosphate, and the tertiary allylic isomer, (3R)-nerolidyl diphosphate (3), using both the native fungal and recombinant enzymes. A series of farnesyl diphosphate analogs, 15, 16, 20, 7, 8, and 9, was also tested as inhibitors of trichodiene synthase. 10-Fluorofarnesyl diphosphate (15) was the most effective competitive inhibitor, with a K1 of 16 nM compared to the Km for 1 of 87 nM, while the ether analog of farnesyl diphosphate, 8, an extremely potent inhibitor of squalene synthase, showed only modest inhibition of trichodiene synthase, with a K1/Km of 70.

Binding, Competitive↗