Nucleic Acids Research, Vol. 18, No. I

Primary structure of guinea pig apolipoprotein E Teruhiko Matsushima'*, Godfrey S.Getz1' 2 and Stephen C.Meredith1 Departments of 1Pathology, Biochemistry and 2Molecular Biology, The University of Chicago, Chicago, IL 60637, USA Submitted September 25, 1989

Guinea pig apolipoprotein (apo) E has been cloned and sequenced. The calculated Mr is 32,374, about 2000 less than that of human apoE (human apoE), and in agreement with previous results by SDS-PAGE. The 5' non-coding region is about one and a half

GP HU GP HU

tgs

(49)

gsc

(109)

Stc ctg tgg gct gct ctg gtg gtc acg ctt ctg gca "ga tgc cgg gca

( 169 )

---

---

---

---

---

k

v

*

q

a

ga gag

GP HU

ccccgc gg agg at ctc tga acc tat ttc-tgc agc ggc 8 cgscg cgc ag gt cg cga Sca "c ctt ggc tca tca *tc sca

ggg agS atg aaS K M K M

acc

ccsg tcsg S

L L

V V

W W

---g v

d *

A

A A

L L

gtg gsa E v E t

cc P

A

P

cag agc gsg cag ccc tgg gag ctt gcg ctg Q S G Q p W E L A L Q S G Q r W E L A L

V V

v

1

T T

1 f

L L

A A

G G

C C

r q

A A

gSag gtg gag gtc cgg gag cca gca gtg tgg E E

V

E E

a.c

cgc R R

p

a

g

v

R

*

1

R

q

tic tgg gat F F

W W

D D

p q

a

v

t

*

tac ctg cgc tgg

Y Y

L L

R R

W W

-199

(211)

W W

14 20

gtg

(271)

V V

34 40

gag ctg ctg tct gac cag gtg cag gag gag ctg ctc agc aac cag gtc acc cag T E L E E L L L S d V Q Q S n Q V Q . 0Q V O E E L L 0 V T O E L S a L

(331) 54

(391)

GP HU

acg ctg ctg att gaSg gac acc atg aag gaggtg ags gcc tac aag gcg gag ctg gag aSg a E L E k 1 d T M K E v K A Y K t L i * A 1 K K E L E0 d e T K E Y s M L m r a Saa gac acg gaggct cgg ctggcc aa gags ctg cag gcg gcS T A R a K E L Q A A d L k E K E L Q A A A R E T L s r *

(451)

GP HU

gaa cta gga g * L t q L

cag GP HU

Q Q

cc

T T

ccggtg gca P P

V V

A A

caS gca cgg ctg ggg gca gac atg GP HU

Q Q

A A

R R

L L

cg cc gag gtScg GP HU

GP HU

GP HU

E E

V V

Q Q

A A

A A

G G

M M

D D

M M

gctggcc G Q L G Q L

gagggicgc E E

* d

V V

R R

c gagS tc S S

a

t

E E

g

GP HU

E E

GP HU GP

HfU

R R

gcg a v

cgc R R

ctc L L

acc t a

tcg S S

cac

H H

94 100

(511) 114 120

(571) 134 140

gat gaa ctg cag ag cgt atg act . L Q K R m A D d K R 1 A Q D L

(631) 154 160

gtg tac *ag gcc gga gc cac gag ggc gcc SgS cgt

Sat

gtg agc gcc atc cga gag cgc S A I R R E v 1 A I R E R S

(691)

acc agc A s

(736)

V V

Y Y

k q

A A

G G

L L

G G

s

p

L L

i v

cag cca ctg cU gag Q P L Q E Q P L Q E

A A

q r

E E

G G

A A

E E

R R

v m

v a

E E

Q Q

G G

R R

1

v

q r

A A

cgc gca cac &cc tgg ggtgag R Q Q A W G E Q A W G E R A

G G

---

---

---

---

---

ctg

I

I t

I

I

I

v

L LI

a

a

cag atg cgc ggg cgg q R m R S a 1 R R r

A

g

ctg gag aag 1

m

E E

k a

ggc aSgc cag gcg cgc gac cgc ctg gag gag gtg cgt gaa cag atg gag gag gtg cgc V E Q V S q a a r m E R E * G R D R L V Q a E V R S R d E k E v D L G r t R

stc as gtg gSag gag GP HU

L L

ctg tcg cag tac cgc aSc a a Q Y R L g R L v Q y

74 80

ccg cgc asg atg cgcacgcgg ctg cag cgc gac atc i q R D p R K a X R R L I R K K R L 1 R D A 1 R

gtg GP HU

R

g ctc csc

ctg ggg tcc ctgs ta Sgaa cag ggc cgt ctt caa gc GP HU

aac cgc n R

60

K K

v

1

E E

E E

c --gaa cag-.

Q

Q

A A

* q

* q

I i

* r

I 1

* q

* a

E E

gcc ttc cag A F Q A F Q

gcg cgc A

A

R R

ctc aag agc tgg ttc gag ccc atg atg Saa gac atg cgg cgc cag tgg gcc gaa ctc atc W A K F M r R Q L i D S W E P m m E * L g K 1 q R W A S W F M Q L v D E P v E L

cag aag gtg cag Q q K V K V Q *

Stg gct v a

A A

gig V V

ggc gcc agc acc tcs G G

a

t

S S

t a

a a

---

gct ccc agc cag gag ccc tga

* p

v

PF P

S S

q d

*

p555

n

h

gca gcc gc ttc cgc ccc acc cca ccc act gga gtc cct cag cca gca gga agc acc agg ctc tgt acc cac ccc cgc ctg ctc ctg gag ggc tct agt tia taa aa tts gaa aa ttt gcc gag cct cgc - (Poly A)

174 180 189 200

(796) 209 220

(856) 229 240

(895) 242 260

(955) 262

280

1012) 280 299

1072) 1132) 1144

Sequence of cDNA clone pGE131, with deduced amino acid sequence. For comparison, the amino acid sequence of human (HU) apoE (1, 2) is aligned with that of the guinea pig (GP). Residues where the human and guinea pig sequences agree are given in upper case letters; other residues in lower case. An asterisk (*) is a deletion of an amino acid; deletions in the cDNA are represented by three dashes (---). Peptide synthesized is underlined; receptor binding region is in boldface. Termination codon is represented by ***. Insert into the 5' untranslated region of guinea pig cDNA is in boldface.

*Present address: First Department of

202

Internal Medicine, University

times as long as the 115 nucleotide sequence of the rat and human cDNAs, with a unique 44 nucleotide sequence inserted between two conserved regions. The 3' non-coding region of the cDNA has 132 nucleotides and a poly(A) tail. A polyadenylation signal AATAAA is found 32 nucleotides upstream of the poly(A) tail. Compared to human apoE, guinea pig apoE has four small clusters of deletions: the N-terminal six amino acids of human apoE, and three deletions near the C-terminus, i.e., between residues 186 and 187 (human apoE residues 193- 197 missing), between residues 236 and 237 (human apoE residues 248 -254 missing), and between residues 274 and 275 (human apoE residue 293 missing). There is high homology between the deduced amnino acid sequence of guinea pig apoE and those of 6 other species (human (1, 2), baboon (3), cynomolgus (4), rabbit (5), rat (6), and mouse (7)). The regions deleted from guinea pig apoE are the least conserved among these species. The LDL receptor binding region is among the most highly conserved. The most conserved domain, however, is amino acids 34 through 54 in guinea pig apoE (amino acids 40-60 in human apoE). In humans, baboons, mice, and rats, this domain coterminates with the 3' end of the third exon (3, 8- 10). A synthetic peptide of residues 35-55 guinea pig apoE, was found to be amphiphilic in monolayer studies and helical in circular dichroic spectroscopy (minima at 204 and 217.5 nm). The conservation and strategic location in the genome suggests a hitherto unrecognized importance of this domain of apoE. REFERENCES 1. McLean,J.W., Elshourbagy,N.A., Chang,D.J., Mahley,R.W. and Taylor,J.M. (1984) J. Biol. Chem. 259, 6498-6504. 2. Das,H.K., McPherson,J., Bruns,G.A.P., Karathanasis,S.K. and Breslow,J.L. (1985) J. Biol. Chem. 260, 6240-6247. 3. Hixson,J.H., Cox L.A. and Borenstein,S. (1988) Genomics 2, 315-323. 4. Marotti,K.R., Whitted,B.E., Castle,C.K.. Polites,H.G. and Melchior,G.W. (1989) Nucl. Acids Res. 17, 1778. 5. Hao,Q.-I., Yamin,T.-t., Pan,T.-c., Chen,S.-I., Chen,B.-s., Kroon,P.A. and Chao,Y.-S. (1987) Atherosclerosis 66, 125-130. 6. McLean,J.W., Fukazawa,C. and Taylor,J.M. (1983) J. Biol. Chem. 258, 8993 -9000. 7. Rajavashisth,T.B., Kaptein,J.S., Reue,K.L. and Lusis,A.J. (1985) Proc. Natd. Acad. Sci. USA 82, 8085-8089. 8. Feng,W.P., Howlett,G.J. and Schreiber,G. (1986) J. Biol. Chem. 261. 13777-13783. 9. Paik,Y.K., Chang,D.J., Reardon,C.A.,Davies,G.E.. Mahley,R.W. and Taylorj.M. (1985) Proc. Natl. Acad. Sci. USA 82, 3445-3449. 10. HoriuchiK., Tajima,S.. Menju,M. and Yamamoto.A. (1989) J. Biochem. 106, 98- 103.

of Tokyo Hospital, Tokyo, Japan

Primary structure of guinea pig apolipoprotein E.

Nucleic Acids Research, Vol. 18, No. I Primary structure of guinea pig apolipoprotein E Teruhiko Matsushima'*, Godfrey S.Getz1' 2 and Stephen C.Mered...
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