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L. BIBBS ET AL. TABLE 1 Summary of Results from ABRF Peptide Synthesis Research Group Study (continued) Percentage Mass Spectrometry Data Correct Product Major Product Other Lys RP- Correct Compo- Protec- Cleavage Samplea HPLC CE Mass Identity nents tion Biotin Chemistry Instrument Reagent IRBJ 0 0 No 2 Biotinsb Dde Biotin-N- Fmoc/HBTU ACT Modified hydroxysuccinimide MPS 396 Reagent-B 1068 84.3 87.2 Yes � Pbf Dde NHS-LC- Fmoc/PyBOP Perseptive Reagent-K Biotin Pioneer 1374 82.8 83.2 Yes � Ala Dde Succin- Fmoc/HBTU/ AB 431 Reagent-R imidyl D-Biotin HOBt 5130 72.9 82.9 Yes Dde D-Biotin Fmoc/HOBt/ DCC AB 431A Reagent-K 5557 96.1 90 Yes Dde D-Biotin Fmoc/HBTU/ HOBt AB 433 Reagent-K 9818 92.1 92.5 Yes Dde D-Biotin Fmoc/HBTU/ AB 432 Modified HOBt Reagent-B 9453 81.6 81.5 Yes Peptide Fmoc (�)Biotin Boc AB 430 90:10 � 90 HF:Anisole 1340T 96 90 Yes MeOtrt D-Biotin Fmoc/HBTU Rainin Modified Symphony Reagent-K 0775 85.1 84 Yes � Biotinb Mtt (�)Biotin Fmoc/TBTU Rainin Modified Symphony Reagent-B 1524 33.7 63.4 Yes (Biotin � Biotin Mtt Sulfo-NHS- Fmoc/TBTU/ Milligen Reagent-K caproate) caproate LC-Biotin HOBt 9050 1609 74.1 66.1 Yes � Biotinb Mtt D-Biotin Fmoc/HBTU/ HOBt AB 432 Reagent-R 1633 60 55.8 Yes � Biotin Mtt EZ-link Fmoc/HOBt/ AB 431A Reagent-K NHS Biotin DCC 3845 0 0 No � Biotin Mtt NHS-LC- Fmoc Opfp/ Milligen Modified Biotin HOBt 9050 Plus Reagent-B 4209M 7 6.8 No � Biotin Mtt (�)Biotin Fmoc/HBTU ACT Modified 396MBS Reagent-B 7298 81.2 70.8 Yes � Biotinb Mtt D-Biotin Fmoc/HBTU/ HOBt AB 433 Reagent-K 7299 89.9 80.8 Yes Mtt D-Biotin Fmoc/HBTU/ HOBt AB 433 Reagent-K 7347 78 83.5 Yes � Biotinb Mtt (�)Biotin Fmoc/HBTU Perseptive Pioneer Reagent-B aThe samples are grouped according to Lys protection and then listed in numerical order. bDid not use Boc-Ala-OH. cThe extra mass is on the Lys(biotin) by tandem mass spectrometry. �, with and without; CE, capillary electrophoresis; RP-HPLC, reverse-phase high-performance liquid chromatography. 160 JOURNAL OF BIOMOLECULAR TECHNIQUES, VOLUME 11, ISSUE 4, DECEMBER 2000
FIGURE 4 removed using dilute TFA or acetic acid/trifluoroethanol/methylene chloride (AcOH/TFE/DCM). 4 4. Two laboratories used Fmoc-Lys(Aloc)-OH. One of these samples contained multiple components, with the product of interest at only 12.5%. For this synthesis, the Aloc group must be selectively removed using Pd(0),Bu 3 SnH, 5 or Pd(0), HOAc, N-methylmorpholine in chloroform. 6 5. Two laboratories constructed their peptides using t-Boc chemistry. In both of these samples, the FIGURE 5 STRATEGIES FOR SYNTHESIS OF LABELED PEPTIDES Electrospray ionization mass spectrum of a submitted sample of AEGKLRFK(biotin) in which there was a caproyl linker between biotin and the C-terminal Lys.The three charge states detected for this peptide indicated a molecular mass of 1286 daltons. product of interest was the major component by HPLC. Overall, nine of the samples did not have the product of interest as the major component by HPLC. Two types of biotin were used by the participating laboratories: biotin and biotin caproate (b-c). The caproate arm increased the mass of the correct peptide to 1286 daltons (Fig. 4) and caused an extra peak in the amino acid analysis corresponding to the Amino acid analysis of a peptide constructed with a caproyl linker to biotin. An extra peak was observed at 28.6 minutes between Met and Lys. JOURNAL OF BIOMOLECULAR TECHNIQUES, VOLUME 11, ISSUE 4, DECEMBER 2000 161
Page 1 and 2: VOLUME 11, ISSUE 4, DECEMBER 2000 C
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Page 5 and 6: CORPORATE SPONSORSHIP ABRF corporat
Page 7 and 8: Implementation of Automation in a S
Page 9 and 10: 0.8 pmol/�L with 5 �L per prime
Page 11 and 12: RESEARCH GROUP REPORTS Strategies f
Page 13 and 14: Matrix-Assisted Laser Desorption Io
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Page 19 and 20: FIGURE 8 FIGURE 9 Tandem mass spect
Page 21 and 22: REFERENCES 1. Angeletti RH, Bonewal
Page 23 and 24: shared research equipment. When ask
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Page 27 and 28: Survey Respondents’ Priorities fo
Page 29 and 30: FIGURE 9 FIGURE 10 Choices for gove
Page 31 and 32: FIGURE 11 Distribution of NIH Resea
Page 33 and 34: BOOK REVIEWS Amino Acid Analysis Pr
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Page 37 and 38: AMINO ACID COMPOSITION AND SEQUENCE
Page 39 and 40: emove contaminants. The bound prote
Page 41 and 42: 2001 DATES TO REMEMBER Jan 13-18 HP
Page 43 and 44: ABRF 2001 THE NEW BIOLOGY Technolog
Page 45 and 46: P5-M Using high speed data collecti
Page 47 and 48: P13-S Microsatellite analysis using
Page 49 and 50: P21-T Versalinx chemical affinity t
Page 51 and 52: P29-M Protein/DNA technology DSL ve
Page 53 and 54: P37-S Data-directed real-time instr
Page 55 and 56: P45-T Identification of changes in
Page 57 and 58: P53-M Nano-LC ion-trap mass spectro
Page 59 and 60: P61-S Genopure: a novel magnetic be
Page 61 and 62: P69-T Efficient detergent and Cooma
Page 63 and 64: P77-M The utilities of N-terminal s
Page 65 and 66: P85-S Rapid oligosaccharide mapping
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P93-T Analytical strategies for the
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P101-M Implementing surface plasmon
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P109-S Protein molecular communicat
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P117-T New separation medium for th
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P125-M A high sensitivity silver st
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P133-S Application of human cDNA mi
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P141-T Somatic embryony patterns an
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R3-M A current profile of microarra
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S4 A crack in the egg: protein-prot
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T1 Oligonucleotide synthesis chemis
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T9 More than one way to capture a p
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Key to Abstract Numbering Prefixes:
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Neitz, S., P38-M Nelson, J., P116-M
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Admon, Arie, 92 Bibbs, Lisa, 1 Bint
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JBT JOURNAL OF BIOMOLECULAR TECHNIQ
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