Sequences of amino acid residues in bacitracin

114 Pages·2016·5.4 MB·English

Iowa State University Capstones, Theses and
Retrospective Theses and Dissertations
Dissertations
1954
Sequences of amino acid residues in bacitracin
John F. Griffith
Iowa State College
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Part of theBiochemistry Commons
Recommended Citation
Griffith, John F., "Sequences of amino acid residues in bacitracin " (1954).Retrospective Theses and Dissertations. 13086.
https://lib.dr.iastate.edu/rtd/13086
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BMmmmB m AMIMO AOID KSi ctus ii lACifMeii
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Approfets
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I@M of Major Depart®@,nt
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B#an -of /'G-raduat# Gell^«gs
Xowa Sta%® Golleg®
1954
UMI Number: DP12304
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OP 551
r~v 3%
c. 1
Page
IlfEODIIGTIOi 1
mttm Of fiE LifiSAfon 3
• Bacitr&eiii A $
Mmlm aelA ooiiipoiltl©n $
StaMlitr 7
fitrstiom feeimTior 8
fniiftloaal gro'tipi ' 9
Amino gPOttp® 9
Carbcdjl group® 11
The •tttll'Mr 11
Ultraviolet absorption spe®traa 14
loleeular weight 15
Affiiii© ®.eit rtiitee stqueEots 16
Otber Baeitr&0in Peptld## If
Amino it©id eoinpoiitioii. If
UltraTiol@t Absorption spectra 20.
.E3g>lEIMEIfAL 22
Materials 22
ia«itracin 22
Chemicale safl reagenti 22
Bacterial ©ulturet 23
Metliodi 23
Mierobiologieal assay® 23
Otiroiiatofr&ptiio laentifioation© on pap@r 2®
Aeeending teehnique ' 26
OtieeMlng tecimiqtie Zf
Location of aalno aeidfl 27
Tll^73
ill
c-uantitatlfe ©teroMst©gr&pfe|' 't>m®X"30
Study of carboxoid lileskiiig ©©aditione
Sequence study I
Sequence study K
Rate study 1. Partial hydrolysis with
hydrochlorie acit-aoetie aclA
Partial hydrolysii
Blocking treptment®
Complete t^drolysig
Rate Btudy II. fartl&l tiydrolysis witM
hydrochloric aeid-acetie acid
terminal residue study with recovery taaplee
Blocking tr#at»snti
Complete k^di^olysl®
Preparation of desiilfwrized b&citraein
Preparation ©f Raney nickel catalyst
Hydrogsnolysie of bacitracin
Qualitative analytis of product
Stqmenee study of detulfurized bacitracin
Blocking trt&tatntfi
Complete hydrolysis
Extraction ©f thlohydantoini ani.
pheny1thiohyd&ntoins
Alkaline l^ydrolysiB of extrseti
Perfowi© acid oxidation ©f bacitracin A
Beqmum study of oxidised bacitracin A
l£S:0Lfi
Nlnhydrin Color Yield®
Carboxoid Blocking le&ction Condition#
Bacitracin Sequenee Studies
Bate Studies
feroinal Residue .Stuiiee
Bacitracin
Desulfurieed bacitracin
Oxidized bacitracin A
Ir
SISCUSSIOH
Carboxoid Bl®©kifig Itaetlou- Gotttitlsnp
Sequence Stwfilts laeltratla
Rate Studies
Terminal ReBidue Stttti®&
Beeitracin
Desulfurized 'bacitraein
OaiitiasA feaeltr-scin 1
SffiEftBX ASB eiMGLCSlOSS
LIfllAfWg i'lfi©
MWOMmmmnm
1
mmommm
leesnt years have produced many variation® and a ftw
Iffiproveasenti In the methods for anino acid reildu© tequtne®
analysis ©f peptldts. the us© ©f 2,^-dinltrofluorobenE©Et
hy Sangtr and alk&lln® phtnyl liothlocyanat# by Edii&n art two
txa»t)l©s of a«lii©id blocking reagenti whloh h&ve found wld«
application. Most of the »®thodg of Bt<|uenee analyels are
of a qualitative nature, used in eonjunotion with iuch tech-
ni«|u@e a.B paper and colu»n chroaat©gra|>hy, paper el@etro-
phortsle, and counter current dletrlhutlon. B©eauee th©e®
art qualitative aethods, thi peptide under eonslderatlon must
h© hoaogeneous, or at least free of other peptide material,
so that other substancte present cannot eaus« misleading
rtsulti. Unfortunately few, if any, of the naturally occur­
ring peptide# are hoaogeneous» so that ofttn laborious puri­
fication proceduree mufit be carried out Cs.g., oxytocin).
In spite of thle, many natural peptides h&ve been purified,
and soae, fluch am ©•raalcldln S, insulin, tyrocldln, oxytocin,
and vasopretiln, have had their amino acid rtsldu® tequences
elucidated.
It has become clear that ftw of the naturally occurring
peptide® have a einple, straight chain gtructure. iever&l
have been shown to be of a cyclic nature, ©tpeciftlly those
2
haTing aatifeletlo Bropertits j. somt eontaln non-peptia®
aoletle#, fhe preeenee of such cyolic arrangenenta of resi­
dues &M ©ther unorthodox linkages, togsther with tht diffi­
culty iii separating and purifying sliiilar peptide® has
suggested that a quantitatif© approach to structure analysis
might perait structure atteralnation of « com|>l« peptid#
which ie the »«.Jor coaponent of a oixture.
fh© antihiotic, Mcltraoin, is such a mixture of pep­
tide®. It ii a faoily of at Itaet six «i»ilar peptides with
& oajor 0onpon®nt, haeitraein 1, providing woat of the anti-
•felotic sctlTity. fht InTeetigatlon upon which this thesle
ie based I0 eonetrned with dtteraining tht tequence of amino
acid reslduts in feaeltraoin toy application of QuantltatlT©
aethodi to the crude aaterial.