First of all, it is difficult to debug what you posted since the code you posted has only 1 row in the HList widget.
Second, I modified it a bit to add a second duplicate row, removed the $main references and it works fine for me.

#!/usr/bin/perl -w
use strict vars;
use Tk;
require Tk::HList;

my $helix_lf = MainWindow->new();
my $helix_result_list=$helix_lf->Scrolled('HList',
              -scrollbars => 'oe',
              -width => 50,
              -height => 20,
              -header => 1,
              -columns => 5,
                -selectbackground=>'green',
               -command =>\&change_hlist_bg
 )->pack(-expand=>1, -fill=>'both', -padx=>2, -pady=>2);

$helix_result_list->headerCreate( 0, -text => 'Chain');
$helix_result_list->headerCreate( 1, -text => 'Ssqno');
$helix_result_list->headerCreate( 2, -text => 'Esqno');
$helix_result_list->headerCreate( 3, -text => 'Class');
$helix_result_list->headerCreate( 4, -text => 'Sequence');

$helix_result_list->add(helix_result_count);
$helix_result_list->add(helix_result_count1);

$helix_result_list->itemCreate(helix_result_count,0,-text=> "one");
$helix_result_list->itemCreate(helix_result_count,1,-text=> "two");
$helix_result_list->itemCreate(helix_result_count,2,-text=> "three");
$helix_result_list->itemCreate(helix_result_count,3,-text=> "four");
$helix_result_list->itemCreate(helix_result_count,4,-text=>"five");

$helix_result_list->itemCreate(helix_result_count1,0,-text=> "one");
$helix_result_list->itemCreate(helix_result_count1,1,-text=> "two");
$helix_result_list->itemCreate(helix_result_count1,2,-text=> "three");
$helix_result_list->itemCreate(helix_result_count1,3,-text=> "four");
$helix_result_list->itemCreate(helix_result_count1,4,-text=>"five");

MainLoop;
[download]

Hope this helps,
davidj

update:The reason I removed the references to "main" package is because I was getting the following error:

D:\PerlProjects\tests\HList.pl
Entry "" not found at C:/Perl/site/lib/Tk.pm line 228.
[download]

Now, you may need the $main references. I couldn't tell by the code you posted. However, try taking them out and see if it works for you.