The non-fluorescence of 4-fluorotryptophan

Hott, John Lester; Borkman, Raymond F.

doi:10.1042/bj2640297

Cited by 26 publications

(22 citation statements)

References 4 publications

(9 reference statements)

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“…The 4-F-trp and 5-Br-trp analogs, however, had weak emission intensity at room temperature (0.039 and 0.022, respectively, compared to tryptophan); 4-F-trp also showed the largest red shift, to 469 nm, of any of the derivatives. The red shift (compared to tryptophan) in the 4-F-trp emission at 20°C contrasts with a 5-6 nm blue shift seen in the 4-Ftrp phosphorescence spectra collected at 77 K in ethylene glycol glass (6); this suggests that the phosphorescence emission of this analog is sensitive to the chemical and physical properties of the local environment. The emission spectrum of each of the analogs was broadened compared to that of tryptophan.…”

Section: Phosphorescence Emissionmentioning

confidence: 70%

“…These same studies also indicate that 4-F-trp is more photoreactive than tryptophan at room temperature (6). This suggests that the substitution of a fluorine for a hydrogen in the 4 position on tryptophan, while subduing its fluorescence emission, may also enhance triplet state formation and phosphorescence emission.…”

Section: Introductionmentioning

confidence: 70%

“…The replacement of a hydrogen atom with fluorine in the 4 position ( Fig. 1) on tryptophan markedly decreases its fluorescence by over 100-fold (6). Replacement of tryptophan with the nonfluorescent analog 4-fluorotryptophan (4-F-trp)?…”

Section: Introductionmentioning

confidence: 99%

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Room Temperature Phosphorescence from Tryptophan and Halogenated Tryptophan Analogs in Amorphous Sucrose

McCaul

Ludescher

1999

Photochem & Photobiology

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Abstract. Tryptophan phosphorescence lifetime and quantum yield are sensitive to the local environment. The phosphorescence from tryptophan analogs, however, has not been studied. We report here data on the room temperature phosphorescence of tryptophan, 4‐, 5‐ and 6‐fluoro‐DL‐tryptophan (4‐F‐trp, 5‐F‐trp and 6‐F‐trp) and 5‐bromo‐DL‐tryptophan (5‐Br‐trp) embedded in glassy powders of freeze‐dried sucrose. In aqueous solution, the absorption of the analogs was either blue‐shifted (4‐F‐trp), red‐shifted (5‐F‐trp and 5‐Br‐trp) or not shifted (6‐F‐trp) with respect to tryptophan. The phosphorescence emission spectra of all analogs were red‐shifted compared to trp (442 nm) with maxima at 446 nm (5‐F‐trp), 451 mn (6‐F‐trp), 452 nm (5‐Br‐trp) and 469 nm (4‐F‐trp). The 5‐F‐trp and 6‐F‐trp analogs had emission intensities similar to tryptophan (relative quantum yields of 0.68 and 0.91, respectively, compared to tryptophan), while the intensities of the 4‐F and 5‐Br analogs were lower (relative quantum yields of 0.039 and 0.022, respectively). All analogs exhibited complex decay behavior requiring several exponentials for an adequate fit; the average lifetimes were all lower than that of trp (1039 ms). The average lifetimes of the fluorinated analogs (5‐F, 721 ms; 6‐F, 482 ms and 4‐F, 35 ms) scaled approximately with the relative quantum yields while that of 5‐Br (0.53 ms) was significantly lower. Analysis of the individual lifetimes suggested that the fluorinated analogs differ in their sensitivity to environmental interactions, with 5‐F‐ and 6‐F‐trp quenched 1.5‐2‐fold and 4‐F‐trp about 23‐fold more efficiently than tryptophan. The red‐shifted 5‐F‐trypto‐phan analog, which has been incorporated into proteins, may provide an alternative phosphorescence probe for selective phosphorescence detection of a specific protein in a complex mixture.

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Section: Phosphorescence Emissionmentioning

confidence: 70%

Section: Introductionmentioning

confidence: 70%

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Room Temperature Phosphorescence from Tryptophan and Halogenated Tryptophan Analogs in Amorphous Sucrose

McCaul

Ludescher

1999

Photochem & Photobiology

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“…Expression of the CAII-encoding gene is required for the differentiation of myelomonocytic cells into the osteoclast phenotype, this enzyme being highly expressed in the fully differentiated osteoclast (24,25) and its inhibition leading to a decrease in bone resorption both in vitro and in vivo (5,26). We and others (27,28) have previously shown that 1,25(OH)2D3, a potent regulator of osteoclast and macrophage differentiation, can induce a marked increase in CAII mRNA and protein expression in myelomonocytic cells after a 24-hr exposure (7,8).…”

Section: Discussionmentioning

confidence: 99%

1 alpha,25-dihydroxyvitamin D3 regulates the transcription of carbonic anhydrase II mRNA in avian myelomonocytes.

Lomri

Baron

1992

Proc. Natl. Acad. Sci. U.S.A.

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Carbonic anhydrase II (CAII) is highly expressed in the osteoclast, where it is involved in the process of extracellular acidification required for bone resorption. We have previously shown that la,25-dihydroxyvitamin D3 [1,25(OH) (1), and the osteoclast in bone (2, 3). In all these cells, the role of CAII is to generate H+ for acid extrusion by the ATP-driven proton pumps, and HCO-is usually extruded via bicarbonate/chloride exchangers. The importance of CAII in the function of osteoclasts is best demonstrated by the fact that mutation ofthe CAII-encoding gene is associated with osteopetrosis, renal tubular acidosis, and cerebral calcification (4). Furthermore, inhibition of CAII in vitro induces a decrease in bone resorption (5), and CAII activity and levels of expression can be regulated by calciotropic and thyroid hormones (6-9). This regulation includes la,25-dihydroxyvitamin D3 [1,25(OH)2D3], the active metabolite of the steroid hormone vitamin D, which regulates CAII mRNA and protein levels in human and avian mononuclear phagocytes (7,8). The action of steroid hormones involves their high-affinity binding to specific receptors in nuclei, and the regulation of specific gene transcription is mediated by binding of the hormone-receptor complex to steroid-responsive elements present in the promoter regions of the affected genes (10). Although analysis of the CAII gene suggested the possibility that the promoter region contained vitamin D-responsive elements (VDRE) (8,11,12), no change in CAII mRNA levels were observed before 24 hr in the human promonocytic cell line HL-60 (7). The present study was, therefore, done to determine whether 1,25(OH)2D3 increases CAII mRNA by acting at the transcriptional and/or posttranscriptional levels and whether this increase required the transcription and translation of other gene products.Using a transformed myelomonocytic avian cell line (BM2), we found that the regulation of CAII levels by 1,25(OH)2D3 in mononuclear phagocytes occurs mostly via an increase in gene transcription. This effect is independent of de novo protein synthesis, thereby suggesting that the effect does not require the synthesis of other gene products. MATERIALS AND METHODS 4688The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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“…By contrast, the absorbance spectra of another group of tryptophan derivatives, the hydroxytryptophans, are sufficiently red-shifted to allow selective excitation in the presence of tryptophan. In addition, several hydroxytryptophan derivatives have high fluorescence quantum yields (4)(5)(6). These properties suggest the use of hydroxytryptophan derivatives for the generation of SEPs, the intrinsic fluorescence of which can be excited selectively in the presence of other, tryptophan-containing proteins.…”

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confidence: 99%

Spectral enhancement of proteins: biological incorporation and fluorescence characterization of 5-hydroxytryptophan in bacteriophage lambda cI repressor.

Ross

Senear

Waxman

et al. 1992

Proc. Natl. Acad. Sci. U.S.A.

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We have used a tryptophan-requiiing Escherichia coli auxotroph to replace the three tryptophan residues of A cI repressor with 5-hydroxy-L-tryptophan (5-OHTrp). By using a nonleaky promoter, we have achieved >95% replacement of tryptophan in the repressor. We show that the absorbance and fluorescence properties of 5-OHTrp-A cI are clearly distinct from A cI repressor and that the fluorescence of 5-OHTrp-A cI repressor can be observed selectively in the presence of exogenous tryptophan. We also show that the 5-OHTrp-A cI repressor functional properties, as assessed by measurement of binding constants for self-association and for association to operator DNA, and structural properties, as assessed by fluorescence, are indistinguishable from the native repressor. Based on these results, we anticipate that the availability of spectrally enhanced proteins wiil significantly enhance the utility of both fluorescence and phosphorescence spectroscopies to study protein structure and function in complex interacting systems.Tryptophan fluorescence has been used widely to study conformation, function, dynamics, and intermolecular interactions of proteins (1). However, the presence of at least one and often several tryptophans in most proteins severely limits the utility oftryptophan fluorescence for studying one protein species in the company of others. For this reason, many fluorescence studies involving proteins and polypeptides make use of extrinsic probes, such as dansyl or fluorescein. Use of extrinsic probes, however, is complicated by the difficulty of specific placement and the risk that chemical modification can alter the functional and structural properties of the labeled protein.Clearly, it would be to great advantage to incorporate into a protein a fluorophore that is spectrally distinct from tryptophan while having none of the disadvantages of extrinsic probes. In this way, a spectrally enhanced protein (SEP) would be generated that would still have "intrinsic" fluorescence. To be useful for the generation of SEPs, a candidate tryptophan analogue mlist be readily incorporated by biosynthesis into the protein ofinterest, must be fluorescent and have spectral properties distinct from those oftryptophan, and must have little or no effect on the functional and structural properties of the protein into which it has been incorporated.An intriguing possibility for creation of SEPs is suggested by NMR studies in which tryptophan auxotrophs of Escherichia coli were used to incorporate fluorotryptophan derivatives into Salmonella typhimurium histidine binding protein J (2) and rat cellular retinol binding protein (3). The fluorotryptophan derivatives, however, are spectrally very similar to tryptophan. Thus, they do not have features that enhance fluorescence studies. By contrast, the absorbance spectra of another group of tryptophan derivatives, the hydroxytryptophans, are sufficiently red-shifted to allow selective excitation in the presence of tryptophan. In addition, several hydroxytryptophan derivatives have ...

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The non-fluorescence of 4-fluorotryptophan

Cited by 26 publications

References 4 publications

Room Temperature Phosphorescence from Tryptophan and Halogenated Tryptophan Analogs in Amorphous Sucrose

Room Temperature Phosphorescence from Tryptophan and Halogenated Tryptophan Analogs in Amorphous Sucrose

1 alpha,25-dihydroxyvitamin D3 regulates the transcription of carbonic anhydrase II mRNA in avian myelomonocytes.

Spectral enhancement of proteins: biological incorporation and fluorescence characterization of 5-hydroxytryptophan in bacteriophage lambda cI repressor.

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