Genetic encoding of a highly photostable, long lifetime fluorescent amino acid for imaging in mammalian cells.

Acridonylalanine (Acd) is a fluorescent amino acid that is highly photostable, with a high quantum yield and long fluorescence lifetime in water. These properties make it superior to existing genetically encodable fluorescent amino acids for monitoring protein interactions and conformational changes through fluorescence polarization or lifetime experiments, including fluorescence lifetime imaging microscopy (FLIM). Here, we report the genetic incorporation of Acd using engineered pyrrolysine tRNA synthetase (RS) mutants that allow for efficient Acd incorporation in both and mammalian cells.

Access to Faster Eukaryotic Cell Labeling with Encoded Tetrazine Amino Acids.

Labeling of biomolecules in live eukaryotic cells has been limited by low component stability and slow reaction rates. We show that genetically encoded tetrazine amino acids in proteins reach reaction rates of 8 × 10 M s with sTCO reagents, making them the fastest site-specific bioorthogonal labels in eukaryotic systems. We demonstrate that tetrazine amino acids are stable on proteins and are capable of quantitative labeling with sTCO reagents.

Growth Hormone Deficiency Causing Micropenis: Lessons Learned From a Well-Adjusted Adult.

This report of a 46,XY patient born with a micropenis consistent with etiology from isolated congenital growth hormone deficiency is used to (1) raise the question regarding what degree testicular testosterone exposure to the central nervous system during fetal life and early infancy has on the development of male gender identity, regardless of gender of rearing; (2) suggest the obligatory nature of timely full disclosure of medical history; (3) emphasize that virtually all 46,XY infants with functional testes and a micropenis should be initially boys except some with partial androgen insensitivity syndrome; and (4) highlight the sustaining value of a positive long-term relationship with a trusted physician (R.M.B.

Site-Specific Protein Labeling with Tetrazine Amino Acids.

Genetic code expansion is commonly used to introduce bioorthogonal reactive functional groups onto proteins for labeling. In recent years, the inverse electron demand Diels-Alder reaction between tetrazines and strained trans-cyclooctenes has increased in popularity as a bioorthogonal ligation for protein labeling due to its fast reaction rate and high in vivo stability. We provide methods for the facile synthesis of a tetrazine containing amino acid, Tet-v2.

Improving target amino acid selectivity in a permissive aminoacyl tRNA synthetase through counter-selection.

The amino acid acridon-2-ylalanine (Acd) can be a valuable probe of protein dynamics, either alone or as part of a Förster resonance energy transfer (FRET) or photo-induced electron transfer (eT) probe pair. We have previously reported the genetic incorporation of Acd by an aminoacyl tRNA synthetase (RS). However, this RS, developed from a library of permissive RSs, also incorporates N-phenyl-aminophenylalanine (Npf), a trace byproduct of one Acd synthetic route.

Ideal Bioorthogonal Reactions Using A Site-Specifically Encoded Tetrazine Amino Acid.

Bioorthogonal reactions for labeling biomolecules in live cells have been limited by slow reaction rates or low component selectivity and stability. Ideal bioorthogonal reactions with high reaction rates, high selectivity, and high stability would allow for stoichiometric labeling of biomolecules in minutes and eliminate the need to wash out excess labeling reagent. Currently, no general method exists for controlled stoichiometric or substoichiometric labeling of proteins in live cells.

1,2,4-Triazines Are Versatile Bioorthogonal Reagents.

A new class of bioorthogonal reagents, 1,2,4-triazines, is described. These scaffolds are stable in biological media and capable of robust reactivity with trans-cyclooctene (TCO). The enhanced stability of the triazine scaffold enabled its direct use in recombinant protein production.

History of growth hormone therapy.

The first human to receive GH therapy was in 1956; it was of bovine origin and was given for 3 wk for metabolic balance studies revealing no effects. By 1958, three separate laboratories utilizing different extraction methods retrieved hGH from human pituitaries, purified it and used for clinical investigation. By 1959 presumed GHD patients were being given native hGH collected and extracted by various methods.

Nursing leadership from bedside to boardroom: a Gallup national survey of opinion leaders.

Objective: : The purpose of this study was to examine how nursing is viewed by the nation's decision makers and opinion leaders.

Background: : Nurses comprise the largest subgroup of the health and healthcare workforce. The public recognizes nurses' contributions, consistently ranking them highest in honesty and ethics.

2003 American Urological Association Gallup survey: physician practice patterns, cryosurgery/brachytherapy, male infertility, female urology and insurance/professional liability.

Purpose: The Health Policy Survey and Research Committee of the American Urological Association and the Gallup Organization have performed 10 surveys of American urologists since 1992 for the purpose of assessing demographics and practice patterns. The results of the 2003 survey are presented.

Materials And Methods: A random sample of 510 urologists who have completed urological residencies was interviewed by telephone in February 2003.

2001 American Urological Association Gallup Survey: changes in physician practice patterns, satisfaction with urology, and treatment of prostate cancer and erectile dysfunction.

Purpose: The Health Policy Survey and Research Committee of the American Urological Association and the Gallup organization have performed 9 surveys of American urologists since 1992 for the purpose of assessing demographics and practice patterns. The results of the 2001 survey are presented.

Materials And Methods: A random sample of 507 urologists was interviewed in February and March 2001.

1999 American Urological Association Gallup Survey: changes in physician practice patterns, treatment of incontinence and bladder cancer, and impact of managed care.

Purpose: The Health Policy Survey and Research Committee of the American Urological Association and the Gallup Organization have performed a yearly survey of American urologists since 1992 to assess practice patterns. The results of the 1999 survey are presented.

Materials And Methods: A random sample of 503 urologists was interviewed in February and March 1999.

1998 American Urological Association Gallup Survey: changes in physician practice patterns, treatment of ureteral stones and impact of managed care.

Purpose: The American Urological Association first commissioned the Gallup Organization to conduct a study to assess urologist practice patterns in 1992. We present the results of the seventh consecutive Gallup Survey performed for the Association.

Materials And Methods: A random sample of 537 American urologists who completed urological residency and practiced at least 20 hours weekly in 1997 were interviewed by telephone in March 1998.

1997 American Urological Association Gallup survey: Changes in diagnosis and management of prostate cancer and benign prostatic hyperplasia, and other practice trends from 1994 to 1997.

Purpose: The American Urological Association first commissioned the Gallup Organization to conduct a study to assess urologist practice patterns in 1992. We present the results of the 1997 survey, the sixth consecutive Gallup survey performed for the Association.

Materials And Methods: A random sample of 502 American urologists who had completed urological residency and practiced at least 20 hours weekly in 1996 was interviewed by telephone in February and March 1997.

Subspecialization, recruitment and retirement trends of American urologists.

Purpose: Trends of urology workforce, subspecialization, recruitment practices, retirement planning, practice characteristics and managed care impacts in the United States were assessed.

Materials And Methods: In February 1996 the executive interviewing branch of The Gallup Organization selected randomly and interviewed by telephone 507 practicing urologists in the United States who had provided urological patient care for more than 20 hours per week, practiced in 1995 and completed a urological residency program.

Results: Several important issues emerged.

Longitudinal assessment of hormonal and physical alterations during normal puberty in boys. IV: Predictions of adult height by the Bayley-Pinneau, Roche-Wainer-Thissen, and Tanner-Whitehouse methods compared.

To determine how accurately the Roche-Wainer-Thissen (RWT), Tanner-Whitehouse (TW2), and Bayley-Pinneau (BP) prediction models estimated adult height, serial height predictions were made for 23 healthy boys (mean initial age 10.4 ± 1.1 years) every 8 months from 8-15 years of age.

A longitudinal assessment of hormonal and physical alterations during normal puberty in boys. III. The neuroendocrine growth hormone axis during late prepuberty.

Cross-sectional studies demonstrating that physiological GH secretion varies widely among normally and slowly growing children as well as adults have created uncertainty over the definition of normal GH secretion. Furthermore, recent data indicate that the pattern of GH released under identical physiological conditions may be unique for each individual, and suggest that normality may be an individually defined condition. To investigate and develop further this concept and to avoid the confounding effects of fluctuating gonadal steroid hormone levels, we chose as a model normally growing prepubertal boys and performed a longitudinal assessment of spontaneous GH release characteristics (36 24-h GH secretion studies in 9 boys over 9-19 months).

Practice trends of American urologists in the treatment of impotence, incontinence and infertility.

Purpose: Trends of urologist practice patterns in evaluating and treating impotence, incontinence and infertility in the United States were assessed.

Materials And Methods: In July 1995 the executive interviewing branch of the Gallup Organization selected randomly and interviewed by telephone 533 practicing urologists in the United States who had provided urological patient care for more than 20 hours per week, practiced in 1994 and completed a urological residency program.

Results: Treatment of male sexual dysfunction and female urinary incontinence comprises a significant portion of the professional activity of United States urologists.

A longitudinal assessment of hormonal and physical alterations during normal puberty in boys. II. Estrogen levels as determined by an ultrasensitive bioassay.

A limited number of reports of estrogen levels in prepubertal and early pubertal boys have been published because previous estrogen assays have lacked adequate sensitivity to quantitate circulating estrogen concentrations. Development of a new ultrasensitive assay has permitted measurement of estrogen levels in 23 normally growing boys progressing through puberty. Concentrations were measured at approximately 4-month intervals over a 5- to 8-yr period.

A recurring dominant negative mutation causes autosomal dominant growth hormone deficiency--a clinical research center study.

Familial isolated GH deficiency type II (IGHD-II) is an autosomal dominant disorder that has been previously shown in some patients to be caused by heterogeneous GH gene defects that affect GH messenger RNA (mRNA) splicing. We report here our finding of multiple G-->A transitions of the first base of the donor splice site of IVS 3 (+ 1G-->A) in IGHD II subjects from three nonrelated kindreds from Sweden, North America, and South Africa. This + 1G-->A substitution creates an NlaIII site that was used to demonstrate that all affected individuals in all three families were heterozygous for the mutation.

The use--and misuse--of growth hormone.

Recombinant growth hormone is an effective treatment for many children with short stature, but certainly not for all of them. The approach outlined here will help you make the right choices.