A haemoglobin containing only δ-chains

BIOCHEMICAL

Vol. 7, No. 6, 1962

A IIfunuxLoBIN

AND BIOPHYSICAL

RESEARCH COMMUNICATIONS

CONTAINING arm &CHAINS

N. Dance and E.R. Huehns* Department of Biochemistry,

Received

March

contain

a-chains,

Gouttas

et al.,

in vlvo

Huehns et al.,

1955;

(rF) 4

Hunt and Lehmann, 1959; Kekwick and Lehmann, in a number

of individuals

Fessas, 1960;

1960;

(Rigas et al.,

gilvestroni

(Ramot et al.,

et al.,

1960).

1959;

Recently,

abnormal haemoglobin besides haemoglobins H and ttBarttstt

been detected

in two individuals

and this

followed

by starch

obtained

by tryptic

to consist

et al.,

1958)

block electrophoresis. hydrolysis

has

with haemoglobin H disease (Huehns,

haemoglobin has now been isolated

one of them (Case 1, Bingle

it

of haemoglobins which do not

1939) and haemoglobin ttBarttstl

1955; Jones et al.,

has been reported

1962),

occurrence

namely haemoglobin H (0

(Ager and Lehmann, I-958;

a further

College London, England

28, 1962

The simultaneous

1960)

University

from the red cells

of

by column chromatography Analysis

and recombination

of the peptides

experiments

have shown

of 6A2-chains.

solely

Method and Results Haemolysates were prepared from the washed red cells addition

of toluene

and water.

for 18 hours against application

trated .

a O.OlM sodium phosphate buffer

to a column of carboxymethyl

been equilibrated retained

The haemoglobin solution

with

on the column. in vacua with

the same buffer. The eluted

concurrent

cellulose

Beit Memorial Fellow 444

was dialysed

pH 7.0 before which had already

Most of the haemoglobin

haemoglobin solution

dialysis

by the

against

A

was

was concen-

the same buffer

before

Vol. 7, No. 6, 1962

BIOCHEMICAL

being subjected pH 8.6

to starch

and I 0.05.

analysis.

AND BIOPHYSICAL

block electrophoresis

Three haemoglobin zones were resolved

and again subjected

to starch

block

time using a 0.OQ.M sodium phosphate buffer zones were 88613, one of which migrated A.

The haemoglobin

the anode, was eluted the recombination

buffer

by this

and

conthis

Two haemoglobin

the same mobility

as haemo-

zone, which migrated

concentrated.

of this

electrophoresis,

pH 7.0.

with

of the other

and "fingerprintl'

Recombination a.1

in a barbiturate

The pigment of the slowest of these zones was eluted,

centrated

globin

RESEARCH COMMUNICATIONS

nearer to

This haemoglobin was used in

experiments

described.

haemoglobin with haemoglobin aA (Huehns et

1961) at pH 7.0 and 25'C produced one new haemoglobin zone which

on starch

gel electrophoresis

of the tryptic

hydrolysate

migrated of this

peptide

by Baglioni pattern

lysate

followed

(1961) showed that

those peptides

In particular,

of haemoglobin

Ingram and gtretton

peptides

(1961)

called

peptide

of

in the

from the a-chain to those of the

found only in.the

A2T-26 and peptide

were clearly

as des-

present

solely

corresponded

the two peptides

A2,

Comparison

A2.

by chromatography

of haemoglobin A2 which arise

were absent and the remaining bA2-chain.

haemoglobin

haemoglobin with that

haemoglobin A2 by electrophoresis cribed

with

hydroA2a by

visible.

Discussion The small number of peptides shows that chain.

this

haemoglobin consists

identical

only of one type

The absence of the peptides

derived

and A,T-26

indicate

presence of peptides chain.

found in the tryptic

The formation with that

A2a

of a new species with

hydrolysate of polypeptide

from the u-chain that

this

and the A2 is the 6 -

an electrophoretic

of haemoglobin A2 in the recombination A

isolated

haemoglobin with haemoglobin a

contains

a polypeptide

chain with

confirms

that

this

mobility of the newly haemoglobin

the same net charge as the 6A2-chain

of haemoglobin A2. This haemoglobin is therefore

called

haemoglobin

6A2 .

Vol. 7, No. 6, 1962

It

BIOCHEMICAL

is of particular

AND BIOPHYSICAL

interest

that

haemoglobins 6A2 and aA occurred

the recombination

at 25OC ma neutral

other haemoglobina which are known to react these conditions

(Huehns and Beaven, 1962)

type of polypeptide Starch

gel electrophoreais

more rapidly

haemoglobins H, 18Bart's1' and 8A2. Greek Cypriot The isolation the haemolysate

presence of this

and that

might be expected also carry synthesis proportion

that

this

that

of 6A2-chains

identification with

of haemoglobin h4from

haemoglobin H disease and the

it

carry

with

four

in many or all

(Feasas, 1960;

Koler and F&gas, 1961)

then the

to be lower than normal

disease.

Several

other

of haemoglobin A2 in haemoglobin it2 might be Indeed,

several

reports

of a second abnormal haemoglobin

in haemoglobin A disease which resembles haemoglobin 446

it

in these cases the

cases examined here.

suggests that

cases of this

from a

of haemoglobin 6A2, and this

authors have also found a decreased proportion haemoglobin H disease and this

a-chain

haemoglobin A2,

at a normal rate,

are used in the formation

support

haemoglobin II disease would

is assumed that

is proceeding

has been found to be so in all

present

others lend strong

of haemoglobins A end A2 arise

of haemoglobin A2 would be expected

aome h-chains

were found in

in London.

individuals If

three

than haemoglobin A, namely

disease is caused by deficient

haemoglobin 8A2*

with

each of these carries

As dL1 normal adults all

and A, and

other individuals

haemoglobin in three

the a-chains pool.

of a

pH ‘7.4 shows that

These four patients

living

of one individual

commonmetabolic

if

families

and positive

to the hypothesis

and

which al.60 consist

of the haemolysates from the three

abnormal haemoglobins migrating

synthesis

1962)

between haemoglobins "Bart*sl'

shows that

probable

The only haemoglobin aA under

in phosphate buffer

haemoglobin H disease available

three

pH.

chain.

haemoglobin aA2 migrates examination

with

of

are haemoglobin H (Huehna and Shooter,

haemoglobin 9art1s1* single

RESEARCH COMMUNICATIONS

sA2 rather

than

Vol. 7, No. 6, 1962

BIOCHEMKAL

AND BIOPHYSICAL

haemoglobin ~1 have appeared and it further

is possible

examples of the haemoglobin described

RESEARCH COMMUNICATIONS

that

these represent

here.

References Ager, J.A.M. and Lehmann, H., Brit.med.J., 9, 929 (1958) Baglioni, C., Biochim.biophys.Acta, G, 392 (1961) Bingle, J.P., Huehns, E.R. and Prankerd, T.A.J., Brit.med.J.,

(1958)

Fessas, P.,

1043 (1960)

Gouttas,

A.,

(1955)

Proc.7th

Congr.europ.Soc.Haemat.,

Fessas, P., Tsevrenis,

London 1959;

B. and Xefteri,

E.,

1389

ii, part

II,

Sang, 26, 911

Huehns, E.R., 8th European Congress of Haematology, Vienna, 1961, in Haemoglobin-colloquium, p.76 (1962) Huehns, E.R. and Beaven, G.H., Proc.Biochem.Soc., in press. Huehns, E.R., Flynn, F.V., Butler, E.A. and Shooter, E.M., Brit.J. Haemat., 6, 388 (1960) Huehns, E.R. and Shooter, E.M., Nature, 193, 1083 (1962) Huehns, E.R., Shooter, E.M., Dance, N., Beaven, G.H. and Shooter, K.V., Nature, 192, 1057 (1961) Hunt, J.A. and Lehmann, H., Nature, I&, 872 (1959) Ingram, V.M. and Stretton, A.O.W., Nature, 190, 1079 (1961) Jones, R.T., Schroeder, W.A., Balog, J.E. and Vinograd, J.R., J.Amer. chem.Soc., &, 3161 (1959) Kekwick, R.A. and Lehmann, H., Nature, I& 158 (1960) Koler, R.D. and Rigas, D.A., Ann.Hum.Genet., 2, 95 (1961) Ramot, B., Sheba, Ch., Fisher, S., Ager, J.A.M. and Lehmann, H., Brit.med.J., ii, 1228 (1959) Rigas, D.A., Koler, R.D. and Osgood, E.E., Science, 121, 372 (1955) M.,Policlinico (Prat.), 2, Silvestroni, E., Bianco, I. and Muaeolini,

41 (196o).